PT	AU	BA	BE	GP	AF	BF	CA	TI	SO	SE	BS	LA	DT	CT	CY	CL	SP	HO	DE	ID	AB	C1	RP	EM	RI	OI	FU	FX	CR	NR	TC	Z9	U1	U2	PU	PI	PA	SN	EI	BN	J9	JI	PD	PY	VL	IS	PN	SU	SI	MA	BP	EP	AR	DI	D2	EA	PG	WC	SC	GA	UT	PM	OA	HC	HP	DA
J	Bendall, SC; Simonds, EF; Qiu, P; Amir, EAD; Krutzik, PO; Finck, R; Bruggner, RV; Melamed, R; Trejo, A; Ornatsky, OI; Balderas, RS; Plevritis, SK; Sachs, K; Pe'er, D; Tanner, SD; Nolan, GP				Bendall, Sean C.; Simonds, Erin F.; Qiu, Peng; Amir, El-ad D.; Krutzik, Peter O.; Finck, Rachel; Bruggner, Robert V.; Melamed, Rachel; Trejo, Angelica; Ornatsky, Olga I.; Balderas, Robert S.; Plevritis, Sylvia K.; Sachs, Karen; Pe'er, Dana; Tanner, Scott D.; Nolan, Garry P.			Single-Cell Mass Cytometry of Differential Immune and Drug Responses Across a Human Hematopoietic Continuum	SCIENCE			English	Article							TYROSINE KINASE INHIBITOR; FLOW-CYTOMETRY; SIGNALING NETWORKS; QUANTITATIVE-ANALYSIS; MYELOGENOUS LEUKEMIA; STEM-CELLS; IDENTIFICATION; LYMPHOCYTE; DASATINIB; ACTIVATION	Flow cytometry is an essential tool for dissecting the functional complexity of hematopoiesis. We used single-cell "mass cytometry" to examine healthy human bone marrow, measuring 34 parameters simultaneously in single cells (binding of 31 antibodies, viability, DNA content, and relative cell size). The signaling behavior of cell subsets spanning a defined hematopoietic hierarchy was monitored with 18 simultaneous markers of functional signaling states perturbed by a set of ex vivo stimuli and inhibitors. The data set allowed for an algorithmically driven assembly of related cell types defined by surface antigen expression, providing a superimposable map of cell signaling responses in combination with drug inhibition. Visualized in this manner, the analysis revealed previously unappreciated instances of both precise signaling responses that were bounded within conventionally defined cell subsets and more continuous phosphorylation responses that crossed cell population boundaries in unexpected manners yet tracked closely with cellular phenotype. Collectively, such single-cell analyses provide system-wide views of immune signaling in healthy human hematopoiesis, against which drug action and disease can be compared for mechanistic studies and pharmacologic intervention.	[Bendall, Sean C.; Simonds, Erin F.; Krutzik, Peter O.; Finck, Rachel; Bruggner, Robert V.; Trejo, Angelica; Sachs, Karen; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Qiu, Peng; Plevritis, Sylvia K.] Stanford Univ, Dept Radiol, Stanford, CA 94305 USA; [Amir, El-ad D.; Melamed, Rachel; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA; [Ornatsky, Olga I.; Tanner, Scott D.] Univ Toronto, Toronto, ON M5S 3H6, Canada; [Ornatsky, Olga I.; Tanner, Scott D.] DVS Sci, Markham, ON L3R 6E7, Canada; [Balderas, Robert S.] BD Biosci, San Diego, CA 95131 USA; [Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA[Bendall, Sean C.; Simonds, Erin F.; Krutzik, Peter O.; Finck, Rachel; Bruggner, Robert V.; Trejo, Angelica; Sachs, Karen; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Qiu, Peng; Plevritis, Sylvia K.] Stanford Univ, Dept Radiol, Stanford, CA 94305 USA; [Amir, El-ad D.; Melamed, Rachel; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA; [Ornatsky, Olga I.; Tanner, Scott D.] Univ Toronto, Toronto, ON M5S 3H6, Canada; [Ornatsky, Olga I.; Tanner, Scott D.] DVS Sci, Markham, ON L3R 6E7, Canada; [Balderas, Robert S.] BD Biosci, San Diego, CA 95131 USA; [Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Pe'er, Dana/0000-0002-9259-8817; Altan-Bonnet, Gregoire/0000-0002-7283-3162; Simonds, Erin/0000-0002-3497-4861; Melamed, Rachel/0000-0003-3089-9806	Damon Runyon Cancer Research Foundation [DRG-2017-09]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, P01 CA034233, 272200700038C, 1R01CA130826, U54 CA149145, 5U54 CA143907, RB2-01592, PN2 EY018228, NOI-HV-00242]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; California Institute for Regenerative MedicineCalifornia Institute for Regenerative Medicine [RB2-01592, DR-01477]; Genome Canada via the Ontario Genomics Institute for Cancer Research; Ontario Research Fund [ORF-GL2-01-003]; Burroughs Wellcome FundBurroughs Wellcome Fund; Packard Fellowship for Science and Engineering	The authors thank W. J. Fantl for critical reading of the. manuscript. S.C.B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09). This work was supported by NIH grants, U19 AI057229, P01 CA034233, 272200700038C, 1R01CA130826, U54 CA149145, 5U54 CA143907, RB2-01592, PN2 EY018228, NOI-HV-00242, and HEALTH.2010.1.2-1 (European Commission grant to the Sweden Diatools Consortium), as well as California Institute for Regenerative Medicine (DR-01477, RB2-01592) to G.P.N. G.P.N. is supported by an endowed chair from Rachtford and Carlota A. Harris. S.D.T. was supported by Genome Canada via the Ontario Genomics Institute for Cancer Research and by the Ontario Research Fund ORF-GL2-01-003. D.P. holds a Career Award at the Scientific Interface from the Burroughs Wellcome Fund and Packard Fellowship for Science and Engineering. Some antibodies were a gift from Becton Dickinson Biosciences. B.B. is a paid employee of Becton Dickinson, and G.P.N. and P.K. are paid consultants for Becton Dickinson Biosciences. G.P.N. is a member of the Board of Directors and a consultant for DVS Sciences. O.O., G.P.N., and S.T. have equity holdings in DVS Biosciences, and S.T. is an employee of DVS Sciences A patent has been applied for on the SPADE algorithm on behalf of Stanford University. Raw data and SPADE trees can be downloaded open access at www.cytobank.org/nolanlab.	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J	Amir, ED; Davis, KL; Tadmor, MD; Simonds, EF; Levine, JH; Bendall, SC; Shenfeld, DK; Krishnaswamy, S; Nolan, GP; Pe'er, D				Amir, El-ad David; Davis, Kara L.; Tadmor, Michelle D.; Simonds, Erin F.; Levine, Jacob H.; Bendall, Sean C.; Shenfeld, Daniel K.; Krishnaswamy, Smita; Nolan, Garry P.; Pe'er, Dana			viSNE enables visualization of high dimensional single-cell data and reveals phenotypic heterogeneity of leukemia	NATURE BIOTECHNOLOGY			English	Article							MINIMAL RESIDUAL DISEASE; FLOW-CYTOMETRY; MASS CYTOMETRY; STEM-CELLS; PROGENITORS; HIERARCHY; NETWORKS; BIOLOGY; GUIDE	New high-dimensional, single-cell technologies offer unprecedented resolution in the analysis of heterogeneous tissues. However, because these technologies can measure dozens of parameters simultaneously in individual cells, data interpretation can be challenging. Here we present viSNE, a tool that allows one to map high-dimensional cytometry data onto two dimensions, yet conserve the high-dimensional structure of the data. viSNE plots individual cells in a visual similar to a scatter plot, while using all pairwise distances in high dimension to determine each cell's location in the plot. We integrated mass cytometry with viSNE to map healthy and cancerous bone marrow samples. Healthy bone marrow automatically maps into a consistent shape, whereas leukemia samples map into malformed shapes that are distinct from healthy bone marrow and from each other. We also use viSNE and mass cytometry to compare leukemia diagnosis and relapse samples, and to identify a rare leukemia population reminiscent of minimal residual disease. viSNE can be applied to any multi-dimensional single-cell technology.	[Amir, El-ad David; Tadmor, Michelle D.; Levine, Jacob H.; Shenfeld, Daniel K.; Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Columbia Initiat Syst Biol, New York, NY 10027 USA; [Davis, Kara L.; Simonds, Erin F.; Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA[Amir, El-ad David; Tadmor, Michelle D.; Levine, Jacob H.; Shenfeld, Daniel K.; Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Columbia Initiat Syst Biol, New York, NY 10027 USA; [Davis, Kara L.; Simonds, Erin F.; Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA	Pe'er, D (reprint author), Columbia Univ, Dept Biol Sci, Columbia Initiat Syst Biol, New York, NY 10027 USA.	dpeer@biology.columbia.edudpeer@biology.columbia.edu	Krishnaswamy, Smita/D-8089-2016; Nolan, Garry/AAE-7903-2019	Krishnaswamy, Smita/0000-0001-5823-1985; Simonds, Erin/0000-0002-3497-4861; Pe'er, Dana/0000-0002-9259-8817	National Science Foundation CAREER awardNational Science Foundation (NSF) [MCB-1149728]; National Institutes of Health Roadmap InitiativeUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA; NIH Director's New Innovator Award Program [1-DP2-OD002414-01]; National Centers for Biomedical Computing [1U54CA121852-01A1]; Alex's Lemonade Fund Young Investigator Award; St. Baldrick's Foundation Scholar Award; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; Rachford and Carlota A. Harris Endowed Professorship; European Commission HEALTH [2010.1.2-1]; Bill and Melinda Gates FoundationGates Foundation [GF12141-137101];  [U19 AI057229];  [P01 CA034233];  [HHSN272200700038C];  [1R01CA130826];  [CIRM DR1-01477];  [RB2-01592];  [NCI RFA CA 09-011];  [NHLBIHV-10-05(2)]	The authors would like to thank N. Friedman, I. Pe'er and O. Litvin for valuable comments. The authors would also like to thank M. Minden (Princess Margaret Hospital), C. Mullighan, J. Downing and I. Radtke (St. Jude Children's Hospital) for generously providing leukemia samples for mass cytometry analysis. This research was supported by the National Science Foundation CAREER award through grant number MCB-1149728, National Institutes of Health Roadmap Initiative, NIH Director's New Innovator Award Program through grant number 1-DP2-OD002414-01 and National Centers for Biomedical Computing Grant 1U54CA121852-01A1. E.D.A. is a Howard Hughes Medical Institute International Student Research Fellow. K.L.D. is supported by Alex's Lemonade Fund Young Investigator Award and St. Baldrick's Foundation Scholar Award. S.C.B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09). G.P.N. is supported by the Rachford and Carlota A. Harris Endowed Professorship and grants from U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, CIRM DR1-01477 and RB2-01592, NCI RFA CA 09-011, NHLBIHV-10-05(2), European Commission HEALTH. 2010.1.2-1, and the Bill and Melinda Gates Foundation (GF12141-137101). D.P. holds a Career Award at the Scientific Interface from the Burroughs Wellcome Fund and Packard Fellowship for Science and Engineering.	Ascoli GA, 2008, NAT REV NEUROSCI, V9, P557, DOI 10.1038/nrn2402; Bandura DR, 2009, ANAL CHEM, V81, P6813, DOI 10.1021/ac901049w; Bendall SC, 2012, NAT BIOTECHNOL, V30, P639, DOI 10.1038/nbt.2283; Bendall SC, 2012, TRENDS IMMUNOL, V33, P323, DOI 10.1016/j.it.2012.02.010; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Benoist C, 2011, SCIENCE, V332, P677, DOI 10.1126/science.1206351; Blondel VD, 2008, J STAT MECH-THEORY E, DOI 10.1088/1742-5468/2008/10/P10008; Bodenmiller B, 2012, NAT BIOTECHNOL, V30, P858, DOI 10.1038/nbt.2317; Borowitz MJ, 2008, BLOOD, V111, P5477, DOI 10.1182/blood-2008-01-132837; Campana D, 2008, BRIT J HAEMATOL, V143, P481, DOI 10.1111/j.1365-2141.2008.07350.x; Cornett DS, 2007, NAT METHODS, V4, P828, DOI 10.1038/NMETH1094; Dalerba P, 2011, NAT BIOTECHNOL, V29, P1120, DOI 10.1038/nbt.2038; Herzenberg LA, 2006, NAT IMMUNOL, V7, P681, DOI 10.1038/ni0706-681; Irish JM, 2004, CELL, V118, P217, DOI 10.1016/j.cell.2004.06.028; Kondo M, 2003, ANNU REV IMMUNOL, V21, P759, DOI 10.1146/annurev.immunol.21.120601.141007; Kotecha N, 2010, CURR PROTOC CYTOM, V10, P10, DOI [10.1002/0471142956.cy1017s53, DOI 10.1002/0471142956.CY1017S53]; Loken MR, 2012, BLOOD, V120, P1581, DOI 10.1182/blood-2012-02-408336; Lubeck E, 2012, NAT METHODS, V9, P743, DOI [10.1038/NMETH.2069, 10.1038/nmeth.2069]; Majeti R, 2007, CELL STEM CELL, V1, P635, DOI 10.1016/j.stem.2007.10.001; Mercer J, 2012, CELL REP, V2, P1036, DOI 10.1016/j.celrep.2012.09.003; O'Brien CA, 2009, SEMIN RADIAT ONCOL, V19, P71, DOI 10.1016/j.semradonc.2008.11.001; Ossenkoppele GJ, 2011, BRIT J HAEMATOL, V153, P421, DOI 10.1111/j.1365-2141.2011.08595.x; Ozsolak F, 2011, NAT REV GENET, V12, P87, DOI 10.1038/nrg2934; Pyne S, 2009, P NATL ACAD SCI USA, V106, P8519, DOI 10.1073/pnas.0903028106; Qian Y, 2010, FRONT ENVIRON SCI EN, V4, P1, DOI 10.1007/s11783-010-1000-9; Qiu P, 2011, NAT BIOTECHNOL, V29, P886, DOI 10.1038/nbt.1991; Sachs K, 2005, SCIENCE, V308, P523, DOI 10.1126/science.1105809; Tarnok A, 2010, CYTOM PART A, V77A, P6, DOI 10.1002/cyto.a.20844; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; van der Maaten LJP, 2009, 2009005 TICCTR TILB, V10, P1, DOI DOI 10.1080/13506280444000102; van Lochem EG, 2004, CYTOM PART B-CLIN CY, V60B, P1, DOI 10.1002/cyto.b.20008; Wakita S, 2013, LEUKEMIA, V27, P1044, DOI 10.1038/leu.2012.317; Wang Z, 2009, NAT REV GENET, V10, P57, DOI 10.1038/nrg2484	33	716	721	16	120	NATURE PUBLISHING GROUP	NEW YORK	75 VARICK ST, 9TH FLR, NEW YORK, NY 10013-1917 USA	1087-0156	1546-1696		NAT BIOTECHNOL	Nat. Biotechnol.	JUN	2013	31	6					545	+		10.1038/nbt.2594			10	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	160IO	WOS:000320113200027	23685480	Green Accepted			2020-07-01	
J	Giesen, C; Wang, HAO; Schapiro, D; Zivanovic, N; Jacobs, A; Hattendorf, B; Schuffler, PJ; Grolimund, D; Buhmann, JM; Brandt, S; Varga, Z; Wild, PJ; Gunther, D; Bodenmiller, B				Giesen, Charlotte; Wang, Hao A. O.; Schapiro, Denis; Zivanovic, Nevena; Jacobs, Andrea; Hattendorf, Bodo; Schueffler, Peter J.; Grolimund, Daniel; Buhmann, Joachim M.; Brandt, Simone; Varga, Zsuzsanna; Wild, Peter J.; Guenther, Detlef; Bodenmiller, Bernd			Highly multiplexed imaging of tumor tissues with subcellular resolution by mass cytometry	NATURE METHODS			English	Article							SPECTROMETRY; CELLS; HETEROGENEITY; TARGET	Mass cytometry enables high-dimensional, single-cell analysis of cell type and state. In mass cytometry, rare earth metals are used as reporters on antibodies. Analysis of metal abundances using the mass cytometer allows determination of marker expression in individual cells. Mass cytometry has previously been applied only to cell suspensions. To gain spatial information, we have coupled immunohistochemical and immunocytochemical methods with high-resolution laser ablation to CyTOF mass cytometry. This approach enables the simultaneous imaging of 32 proteins and protein modifications at subcellular resolution; with the availability of additional isotopes, measurement of over 100 markers will be possible. We applied imaging mass cytometry to human breast cancer samples, allowing delineation of cell subpopulations and cell-cell interactions and highlighting tumor heterogeneity. Imaging mass cytometry complements existing imaging approaches. It will enable basic studies of tissue heterogeneity and function and support the transition of medicine toward individualized molecularly targeted diagnosis and therapies.	[Giesen, Charlotte; Schapiro, Denis; Zivanovic, Nevena; Jacobs, Andrea; Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Wang, Hao A. O.; Hattendorf, Bodo; Guenther, Detlef] Swiss Fed Inst Technol, Dept Chem, Zurich, Switzerland; [Wang, Hao A. O.; Grolimund, Daniel] Paul Scherrer Inst, Swiss Light Source, Villigen, Switzerland; [Schapiro, Denis] Swiss Fed Inst Technol, Life Sci Zurich Grad Sch, Syst Biol PhD Program, Zurich, Switzerland; [Schapiro, Denis; Zivanovic, Nevena] Univ Zurich, Zurich, Switzerland; [Zivanovic, Nevena] Swiss Fed Inst Technol, Life Sci Zurich Grad Sch, Mol Life Sci PhD Program, Zurich, Switzerland; [Schueffler, Peter J.; Buhmann, Joachim M.] Swiss Fed Inst Technol, Dept Comp Sci, Zurich, Switzerland; [Brandt, Simone; Varga, Zsuzsanna; Wild, Peter J.] Univ Zurich Hosp, Inst Surg Pathol, CH-8091 Zurich, Switzerland[Giesen, Charlotte; Schapiro, Denis; Zivanovic, Nevena; Jacobs, Andrea; Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Wang, Hao A. O.; Hattendorf, Bodo; Guenther, Detlef] Swiss Fed Inst Technol, Dept Chem, Zurich, Switzerland; [Wang, Hao A. O.; Grolimund, Daniel] Paul Scherrer Inst, Swiss Light Source, Villigen, Switzerland; [Schapiro, Denis] Swiss Fed Inst Technol, Life Sci Zurich Grad Sch, Syst Biol PhD Program, Zurich, Switzerland; [Schapiro, Denis; Zivanovic, Nevena] Univ Zurich, Zurich, Switzerland; [Zivanovic, Nevena] Swiss Fed Inst Technol, Life Sci Zurich Grad Sch, Mol Life Sci PhD Program, Zurich, Switzerland; [Schueffler, Peter J.; Buhmann, Joachim M.] Swiss Fed Inst Technol, Dept Comp Sci, Zurich, Switzerland; [Brandt, Simone; Varga, Zsuzsanna; Wild, Peter J.] Univ Zurich Hosp, Inst Surg Pathol, CH-8091 Zurich, Switzerland	Bodenmiller, B (reprint author), Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland.	guenther@inorg.chem.ethz.ch; bernd.bodenmiller@imls.uzh.chguenther@inorg.chem.ethz.ch; bernd.bodenmiller@imls.uzh.ch	Grolimund, Daniel/O-3127-2017; MS-Powder+MS-Surfdiff+Micro-XAS+Super-XAS, SLS/M-1551-2019; Schuffler, Peter/C-1821-2018	Grolimund, Daniel/0000-0001-9721-7940; Schuffler, Peter/0000-0002-1353-8921; Schapiro, Denis/0000-0002-9391-5722; Hattendorf, Bodo/0000-0001-9991-2410	Society in Science; The Branco Weiss Fellowship; Swiss National Science Foundation (SNSF)Swiss National Science Foundation (SNSF) [200021-119779, 31003A-143877]; ETH Zurich Pioneer Fellowship; SystemsX PhosphoNet-PPM; Baugarten Foundation; EU VIGOR++ project [270379]; SNSF R'Equip [316030-139220]; SNSF Assistant Professorship [PP00P3-144874]; Swiss Cancer League; European Research Council (ERC) under the European Union's Seventh Framework ProgrammeEuropean Research Council (ERC) [336921]	We thank M. Storz for preparing the histological slides and the TMA sections; S. Dettwiler, A. Bohnert, A. Fitsche; the entire Trace Element and Micro Analysis group at ETH Zurich for their experimental support and discussions; N. Daga and C. von Mering for their feedback on data analysis; the ETHZ LAC workshop for their support in design and construction of the laser ablation chamber; and the Lehner and Luschnig groups for giving us access to their immunofluorescence microscopes. This work was supported by a Society in Science, The Branco Weiss Fellowship, administered by the ETH Zurich (C.G.); the Swiss National Science Foundation (SNSF) project grants 200021-119779 (H.A.O.W.), 200021- 119779 (D.Gunther), 31003A-143877 (D.Gunther) and 31003A- 143877(B.B.); an ETH Zurich Pioneer Fellowship (H.A.O.W.); the SystemsX PhosphoNet-PPM grant (P.J.W. and B.B.); a Baugarten Foundation grant (SGGP) (P.J.W.); a EU VIGOR++ project FP7/2007-2013, # 270379 (P.J.S. and J. M. B.); an SNSF R'Equip grant 316030-139220 (B. B.); an SNSF Assistant Professorship grant PP00P3- 144874 (B.B.); a Swiss Cancer League grant (B.B.); and funding from the European Research Council (ERC) under the European Union's Seventh Framework Programme (FP7/2007-2013)/ ERC Grant Agreement no. 336921 (B.B.).	Bandura DR, 2009, ANAL CHEM, V81, P6813, DOI 10.1021/ac901049w; Becker JS, 2010, ANAL CHEM, V82, P9528, DOI 10.1021/ac102256q; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Blurry R. 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Methods	APR	2014	11	4					417	+		10.1038/NMETH.2869			9	Biochemical Research Methods	Biochemistry & Molecular Biology	AE1SL	WOS:000333749900024	24584193				2020-07-01	
J	Qiu, P; Simonds, EF; Bendall, SC; Gibbs, KD; Bruggner, RV; Linderman, MD; Sachs, K; Nolan, GP; Plevritis, SK				Qiu, Peng; Simonds, Erin F.; Bendall, Sean C.; Gibbs, Kenneth D., Jr.; Bruggner, Robert V.; Linderman, Michael D.; Sachs, Karen; Nolan, Garry P.; Plevritis, Sylvia K.			Extracting a cellular hierarchy from high-dimensional cytometry data with SPADE	NATURE BIOTECHNOLOGY			English	Article							FLOW-CYTOMETRY; MASS CYTOMETRY; STEM-CELL; IDENTIFICATION; IMMUNE	The ability to analyze multiple single-cell parameters is critical for understanding cellular heterogeneity. Despite recent advances in measurement technology, methods for analyzing high-dimensional single-cell data are often subjective, labor intensive and require prior knowledge of the biological system. To objectively uncover cellular heterogeneity from single-cell measurements, we present a versatile computational approach, spanning-tree progression analysis of density-normalized events (SPADE). We applied SPADE to flow cytometry data of mouse bone marrow and to mass cytometry data of human bone marrow. In both cases, SPADE organized cells in a hierarchy of related phenotypes that partially recapitulated well-described patterns of hematopoiesis. We demonstrate that SPADE is robust to measurement noise and to the choice of cellular markers. SPADE facilitates the analysis of cellular heterogeneity, the identification of cell types and comparison of functional markers in response to perturbations.	[Qiu, Peng; Plevritis, Sylvia K.] Stanford Univ, Dept Radiol, Stanford, CA 94305 USA; [Qiu, Peng] Univ Texas MD Anderson Canc Ctr, Dept Bioinformat & Computat Biol, Houston, TX 77030 USA; [Simonds, Erin F.; Bendall, Sean C.; Gibbs, Kenneth D., Jr.; Bruggner, Robert V.; Sachs, Karen; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Linderman, Michael D.] Stanford Univ, Comp Syst Lab, Stanford, CA 94305 USA[Qiu, Peng; Plevritis, Sylvia K.] Stanford Univ, Dept Radiol, Stanford, CA 94305 USA; [Qiu, Peng] Univ Texas MD Anderson Canc Ctr, Dept Bioinformat & Computat Biol, Houston, TX 77030 USA; [Simonds, Erin F.; Bendall, Sean C.; Gibbs, Kenneth D., Jr.; Bruggner, Robert V.; Sachs, Karen; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Linderman, Michael D.] Stanford Univ, Comp Syst Lab, Stanford, CA 94305 USA	Qiu, P (reprint author), Stanford Univ, Dept Radiol, Stanford, CA 94305 USA.	pqiu@mdanderson.orgpqiu@mdanderson.org	Nolan, Garry/AAE-7903-2019	Simonds, Erin/0000-0002-3497-4861; Gibbs, Kenneth/0000-0002-3532-5396	National Cancer InstituteUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [U56CA112973, U54CA149145]; A Damon Runyon Cancer Research Foundation; National Science FoundationNational Science Foundation (NSF); Stanford DARE; US National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, 5U54 CA143907, RB2-01592, PN2EY018228, N01-HV-00242]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; California Institute for Regenerative MedicineCalifornia Institute for Regenerative Medicine [DR1-01477]	The authors gratefully acknowledge funding from National Cancer Institute Integrative Cancer Biology Program (ICBP), grants U56CA112973 and U54CA149145 to S.K.P. A Damon Runyon Cancer Research Foundation Fellowship supports S.C.B. National Science Foundation Graduate Research Fellowship and Stanford DARE Fellowship support K. D. G. This work is also supported by US National Institutes of Health grants U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, 5U54 CA143907, RB2-01592, PN2EY018228, N01-HV-00242, HEALTH.2010.1.2-1 (European Commission), as well as the California Institute for Regenerative Medicine (DR1-01477) to G.P.N.	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Biotechnol.	OCT	2011	29	10					886	U181		10.1038/nbt.1991			8	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	838FQ	WOS:000296273000015	21964415	Green Accepted			2020-07-01	
J	Maecker, HT; McCoy, JP; Nussenblatt, R				Maecker, Holden T.; McCoy, J. Philip; Nussenblatt, Robert			Standardizing immunophenotyping for the Human Immunology Project	NATURE REVIEWS IMMUNOLOGY			English	Review							B-CELL SUBPOPULATIONS; CD8(+) T-CELLS; FLOW-CYTOMETRY; MASS CYTOMETRY; NK CELLS; EXPRESSION; SUBSETS; LYMPHOCYTES; DIFFERENTIATION; ANTIGENS	The heterogeneity in the healthy human immune system, and the immunological changes that portend various diseases, have been only partially described. Their comprehensive elucidation has been termed the 'Human Immunology Project'. The accurate measurement of variations in the human immune system requires precise and standardized assays to distinguish true biological changes from technical artefacts. Thus, to be successful, the Human Immunology Project will require standardized assays for immunophenotyping humans in health and disease. A major tool in this effort is flow cytometry, which remains highly variable with regard to sample handling, reagents, instrument setup and data analysis. In this Review, we outline the current state of standardization of flow cytometry assays and summarize the steps that are required to enable the Human Immunology Project.	[Maecker, Holden T.] Stanford Univ, Inst Immun Transplantat & Infect, Sch Med, Stanford, CA 94305 USA; [McCoy, J. Philip] NHLBI, NIH, Bethesda, MD 20892 USA; [McCoy, J. Philip; Nussenblatt, Robert] NIH, Ctr Human Immunol Autoimmun & Inflammat, Bethesda, MD 20892 USA; [Nussenblatt, Robert] NEI, NIH, Bethesda, MD 20893 USA[Maecker, Holden T.] Stanford Univ, Inst Immun Transplantat & Infect, Sch Med, Stanford, CA 94305 USA; [McCoy, J. Philip] NHLBI, NIH, Bethesda, MD 20892 USA; [McCoy, J. 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C, 2006, CURR PROTOC CYTOM, V36, P6231; Rogers Susan, 2007, Yale Journal of Biology and Medicine, V80, P165; Sallusto F, 1999, NATURE, V401, P708, DOI 10.1038/44385; Shi LM, 2006, NAT BIOTECHNOL, V24, P1151, DOI 10.1038/nbt1239; Suni Maria A, 2003, BMC Immunol, V4, P9, DOI 10.1186/1471-2172-4-9; Tussey L, 2000, EUR J IMMUNOL, V30, P1823, DOI 10.1002/1521-4141(200007)30:7<1823::AID-IMMU1823>3.0.CO;2-6; Weinberg A, 2009, CLIN VACCINE IMMUNOL, V16, P1176, DOI 10.1128/CVI.00342-08; Willmann K, 2000, J LEUKOCYTE BIOL, V67, P536; Wrammert J, 2008, NATURE, V453, P667, DOI 10.1038/nature06890; ZIEGLERHEITBROCK HWL, 1991, PATHOBIOLOGY, V59, P127, DOI 10.1159/000163629	61	450	465	8	93	NATURE PUBLISHING GROUP	LONDON	MACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLAND	1474-1733	1474-1741		NAT REV IMMUNOL	Nat. Rev. Immunol.	MAR	2012	12	3					191	200		10.1038/nri3158			10	Immunology	Immunology	899CJ	WOS:000300790600013	22343568	Green Accepted			2020-07-01	
J	Ransohoff, RM				Ransohoff, Richard M.			A polarizing question: do M1 and M2 microglia exist?	NATURE NEUROSCIENCE			English	Article							PRONEUROGENIC PHENOTYPE; MACROPHAGE ACTIVATION; ANALYSIS REVEALS; ADULT MICROGLIA; NEURONAL DEATH; MYELOID CELLS; MOUSE; EXPRESSION; FATE; TRANSCRIPTOME	Microglial research has entered a fertile, dynamic phase characterized by novel technologies including two-photon imaging, whole-genome transcriptomic and epigenomic analysis with complementary bioinformatics, unbiased proteomics, cytometry by time of flight (CyTOF; Fluidigm) cytometry, and complex high-content experimental models including slice culture and zebrafish. Against this vivid background of newly emerging data, investigators will encounter in the microglial research literature a body of published work using the terminology of macrophage polarization, most commonly into the M1 and M2 phenotypes. It is the assertion of this opinion piece that microglial polarization has not been established by research findings. Rather, the adoption of this schema was undertaken in an attempt to simplify data interpretation at a time when the ontogeny and functional significance of microglia had not yet been characterized. Now, terminology suggesting established meaningful pathways of microglial polarization hinders rather than aids research progress and should be discarded.	[Ransohoff, Richard M.] Biogen Inc, Neuroimmunol, 14 Cambridge Ctr, Cambridge, MA 02142 USA[Ransohoff, Richard M.] Biogen Inc, Neuroimmunol, 14 Cambridge Ctr, Cambridge, MA 02142 USA	Ransohoff, RM (reprint author), Biogen Inc, Neuroimmunol, 14 Cambridge Ctr, Cambridge, MA 02142 USA.	richard.ransohoff@biogen.comrichard.ransohoff@biogen.com					Adams RA, 2007, J EXP MED, V204, P571, DOI 10.1084/jem.20061931; Ajami B, 2007, NAT NEUROSCI, V10, P1538, DOI 10.1038/nn2014; Ajami B, 2011, NAT NEUROSCI, V14, P1142, DOI 10.1038/nn.2887; ALLIOT F, 1991, P NATL ACAD SCI USA, V88, P1541, DOI 10.1073/pnas.88.4.1541; Alliot F, 1999, DEV BRAIN RES, V117, P145, DOI 10.1016/S0165-3806(99)00113-3; Amit I, 2016, NAT IMMUNOL, V17, P18, DOI 10.1038/ni.3325; Bennett ML, 2016, P NATL ACAD SCI USA, V113, pE1738, DOI 10.1073/pnas.1525528113; Brown GC, 2012, TRENDS BIOCHEM SCI, V37, P325, DOI 10.1016/j.tibs.2012.05.002; Butovsky O, 2014, NAT NEUROSCI, V17, P131, DOI 10.1038/nn.3599; Chen SK, 2010, CELL, V141, P775, DOI 10.1016/j.cell.2010.03.055; Chiu IM, 2013, CELL REP, V4, P385, DOI 10.1016/j.celrep.2013.06.018; Crotti A, 2016, IMMUNITY, V44, P505, DOI 10.1016/j.immuni.2016.02.013; Davalos D, 2005, NAT NEUROSCI, V8, P752, DOI 10.1038/nn1472; Fields RD, 2014, NEUROSCIENTIST, V20, P426, DOI 10.1177/1073858413504465; Gautier EL, 2012, NAT IMMUNOL, V13, P1118, DOI 10.1038/ni.2419; Ginhoux F, 2010, SCIENCE, V330, P841, DOI 10.1126/science.1194637; Goldmann T, 2013, NAT NEUROSCI, V16, P1618, DOI 10.1038/nn.3531; Gosselin D, 2014, CELL, V159, P1327, DOI 10.1016/j.cell.2014.11.023; Grabert K, 2016, NAT NEUROSCI, V19, P504, DOI 10.1038/nn.4222; Hellwig S, 2015, SCI REP-UK, V5, DOI 10.1038/srep14624; Hellwig S, 2013, FRONT CELL NEUROSCI, V7, DOI 10.3389/fncel.2013.00071; Hickman SE, 2013, NAT NEUROSCI, V16, P1896, DOI 10.1038/nn.3554; Holtman IR, 2015, GLIA, V63, P1495, DOI 10.1002/glia.22810; Hoshiko M, 2012, J NEUROSCI, V32, P15106, DOI 10.1523/JNEUROSCI.1167-12.2012; Kan MJ, 2015, J NEUROSCI, V35, P5969, DOI 10.1523/JNEUROSCI.4668-14.2015; Kim CC, 2016, J NEUROINFLAMM, V13, DOI 10.1186/s12974-016-0581-z; Kohman RA, 2012, BRAIN BEHAV IMMUN, V26, P803, DOI 10.1016/j.bbi.2011.10.006; Kumar D, 2016, MOL CELL PROTEOMICS, V15, P329, DOI 10.1074/mcp.M114.047126; Li Y, 2012, DEV CELL, V23, P1189, DOI 10.1016/j.devcel.2012.10.027; Link V.M., 2015, COLD SPRING HARB SYM; Lyons DA, 2015, CSH PERSPECT BIOL, V7, DOI 10.1101/cshperspect.a020586; Martinez Fernando O, 2014, F1000Prime Rep, V6, P13, DOI 10.12703/P6-13; Morganti JM, 2016, PLOS ONE, V11, DOI 10.1371/journal.pone.0148001; Murray PJ, 2014, IMMUNITY, V41, P14, DOI 10.1016/j.immuni.2014.06.008; Neher JJ, 2013, P NATL ACAD SCI USA, V110, pE4098, DOI 10.1073/pnas.1308679110; Nimmerjahn A, 2005, SCIENCE, V308, P1314, DOI 10.1126/science.1110647; Obermeier B, 2013, NAT MED, V19, P1584, DOI 10.1038/nm.3407; Olah M, 2012, GLIA, V60, P96, DOI 10.1002/glia.21251; Oosterhof N, 2015, GLIA, V63, P719, DOI 10.1002/glia.22780; Panatier A, 2012, DEV CELL, V23, P1125, DOI 10.1016/j.devcel.2012.11.015; Paolicelli RC, 2011, NEURON GLIA BIOL, V7, P77, DOI 10.1017/S1740925X12000105; Paolicelli RC, 2011, SCIENCE, V333, P1456, DOI 10.1126/science.1202529; Parkhurst CN, 2013, CELL, V155, P1596, DOI 10.1016/j.cell.2013.11.030; Perdiguero EG, 2013, GLIA, V61, P112, DOI 10.1002/glia.22393; Pont-Lezica L, 2014, EUR J NEUROSCI, V39, P1551, DOI 10.1111/ejn.12508; Ransohoff RM, 2010, NATURE, V468, P253, DOI 10.1038/nature09615; Romanoski CE, 2015, TRENDS IMMUNOL, V36, P507, DOI 10.1016/j.it.2015.07.006; Schafer DP, 2012, NEURON, V74, P691, DOI 10.1016/j.neuron.2012.03.026; Schulz C, 2012, SCIENCE, V336, P86, DOI 10.1126/science.1219179; Sekar A, 2016, NATURE, V530, P177, DOI 10.1038/nature16549; Squarzoni P, 2014, CELL REP, V8, P1271, DOI 10.1016/j.celrep.2014.07.042; Stevens B, 2007, CELL, V131, P1164, DOI 10.1016/j.cell.2007.10.036; Sun SY, 2015, P NATL ACAD SCI USA, V112, pE6993, DOI 10.1073/pnas.1520639112; Thored P, 2009, GLIA, V57, P835, DOI 10.1002/glia.20810; Tremblay ME, 2015, FRONT NEUROANAT, V9, DOI 10.3389/fnana.2015.00045; Tremblay ME, 2011, J NEUROSCI, V31, P16064, DOI 10.1523/JNEUROSCI.4158-11.2011; Ueno M, 2013, NAT NEUROSCI, V16, P543, DOI 10.1038/nn.3358; Viader A, 2016, ELIFE, V5, DOI 10.7554/eLife.12345; Wake H, 2009, J NEUROSCI, V29, P3974, DOI 10.1523/JNEUROSCI.4363-08.2009; Wakselman S, 2008, J NEUROSCI, V28, P8138, DOI 10.1523/JNEUROSCI.1006-08.2008; Wes PD, 2016, GLIA, V64, P197, DOI 10.1002/glia.22866; Xu J, 2015, DEV CELL, V34, P632, DOI 10.1016/j.devcel.2015.08.018; Xue J, 2014, IMMUNITY, V40, P274, DOI 10.1016/j.immuni.2014.01.006; Yamasaki R, 2014, J EXP MED, V211, P1533, DOI 10.1084/jem.20132477; Zhang Y, 2014, J NEUROSCI, V34, P11929, DOI [10.1523/JNEUROSCI.1860-14.2014, 10.11772/j.issn.1001-9081.2014.07.1929]	65	435	442	19	166	NATURE PUBLISHING GROUP	NEW YORK	75 VARICK ST, 9TH FLR, NEW YORK, NY 10013-1917 USA	1097-6256	1546-1726		NAT NEUROSCI	Nat. Neurosci.	AUG	2016	19	8					987	991		10.1038/nn.4338			5	Neurosciences	Neurosciences & Neurology	DS4TB	WOS:000380773200004	27459405				2020-07-01	
J	Levine, JH; Simonds, EF; Bendall, SC; Davis, KL; Amir, EAD; Tadmor, MD; Litvin, O; Fienberg, HG; Jager, A; Zunder, ER; Finck, R; Gedman, AL; Radtke, I; Downing, JR; Pe'er, D; Nolan, GP				Levine, Jacob H.; Simonds, Erin F.; Bendall, Sean C.; Davis, Kara L.; Amir, El-ad D.; Tadmor, Michelle D.; Litvin, Oren; Fienberg, Harris G.; Jager, Astraea; Zunder, Eli R.; Finck, Rachel; Gedman, Amanda L.; Radtke, Ina; Downing, James R.; Pe'er, Dana; Nolan, Garry P.			Data-Driven Phenotypic Dissection of AML Reveals Progenitor-like Cells that Correlate with Prognosis	CELL			English	Article							ACUTE MYELOID-LEUKEMIA; MASS CYTOMETRY; HETEROGENEITY; CANCER; DECONVOLUTION	Acute myeloid leukemia (AML) manifests as phenotypically and functionally diverse cells, often within the same patient. Intratumor phenotypic and functional heterogeneity have been linked primarily by physical sorting experiments, which assume that functionally distinct subpopulations can be prospectively isolated by surface phenotypes. This assumption has proven problematic, and we therefore developed a data-driven approach. Using mass cytometry, we profiled surface and intracellular signaling proteins simultaneously in millions of healthy and leukemic cells. We developed PhenoGraph, which algorithmically defines phenotypes in high-dimensional single-cell data. PhenoGraph revealed that the surface phenotypes of leukemic blasts do not necessarily reflect their intracellular state. Using hematopoietic progenitors, we defined a signaling-based measure of cellular phenotype, which led to isolation of a gene expression signature that was predictive of survival in independent cohorts. This study presents new methods for large-scale analysis of single-cell heterogeneity and demonstrates their utility, yielding insights into AML pathophysiology.	[Levine, Jacob H.; Amir, El-ad D.; Tadmor, Michelle D.; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA; [Levine, Jacob H.; Amir, El-ad D.; Tadmor, Michelle D.; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Syst Biol, New York, NY 10027 USA; [Simonds, Erin F.; Davis, Kara L.; Fienberg, Harris G.; Jager, Astraea; Zunder, Eli R.; Finck, Rachel; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Bendall, Sean C.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Gedman, Amanda L.; Radtke, Ina; Downing, James R.] St Jude Childrens Res Hosp, Dept Pathol, Memphis, TN 38105 USA[Levine, Jacob H.; Amir, El-ad D.; Tadmor, Michelle D.; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA; [Levine, Jacob H.; Amir, El-ad D.; Tadmor, Michelle D.; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Syst Biol, New York, NY 10027 USA; [Simonds, Erin F.; Davis, Kara L.; Fienberg, Harris G.; Jager, Astraea; Zunder, Eli R.; Finck, Rachel; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Bendall, Sean C.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Gedman, Amanda L.; Radtke, Ina; Downing, James R.] St Jude Childrens Res Hosp, Dept Pathol, Memphis, TN 38105 USA	Pe'er, D (reprint author), Columbia Univ, Dept Biol Sci, New York, NY 10027 USA.	dpeer@biology.columbia.edu; gnolan@stanford.edudpeer@biology.columbia.edu; gnolan@stanford.edu	Nolan, Garry/AAE-7903-2019; Downing, James R./N-8102-2018	Pe'er, Dana/0000-0002-9259-8817; Simonds, Erin/0000-0002-3497-4861	DRCRF Fellowships [DRG 2190-14, DRG-2017-09]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [1R00-GM104148, DP1-HD084071, DP2-OD002414, R01-CA164729 U54-CA121852, 1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, P01 CA034233, U19 AI057229, U54CA149145]; Stand Up To Cancer Phillip A. Sharp Award [SU2C-AACR-PS04]; CIRMCalifornia Institute for Regenerative Medicine [DR1-01477, RB2-01592]; ECEuropean Commission Joint Research CentreEuropean Community (EC) [HEALTH.2010.1.2-1]; US FDAUnited States Department of Health & Human Services [HHSF223201210194C]; US DODUnited States Department of Defense [W81XWH-12-1-0591]; Entertainment Industry Foundation; Rachford and Carlota Harris Endowed Professorship; Packard Fellowship for Science and Engineering	We thank G. Behbehani, W. Fantl, B.J. Chen, and L. Zelnik for helpful discussion. E.F.S. and S.C.B. are supported by DRCRF Fellowships (DRG 2190-14 & DRG-2017-09) and NIH 1R00-GM104148 to S.C.B. Grants from NIH (DP1-HD084071, DP2-OD002414, R01-CA164729 U54-CA121852), Stand Up To Cancer Phillip A. Sharp Award SU2C-AACR-PS04 and Packard Fellowship for Science and Engineering supported D.P. Grants from NIH (1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, P01 CA034233, U19 AI057229 and U54CA149145), CIRM (DR1-01477 and RB2-01592), EC (HEALTH.2010.1.2-1), US FDA (HHSF223201210194C), US DOD (W81XWH-12-1-0591), the Entertainment Industry Foundation, and the Rachford and Carlota Harris Endowed Professorship supported G.P.N. G.P.N., S.C.B, H.G.F., and E.F.S. have a personal financial interest in the company Fluidigm, the manufacturer of the mass cytometer used in this manuscript.	Aghaeepour N, 2013, NAT METHODS, V10, P228, DOI [10.1038/NMETH.2365, 10.1038/nmeth.2365]; Akavia UD, 2010, CELL, V143, P1005, DOI 10.1016/j.cell.2010.11.013; Bendall SC, 2014, CELL, V157, P714, DOI 10.1016/j.cell.2014.04.005; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Blondel VD, 2008, J STAT MECH-THEORY E, DOI 10.1088/1742-5468/2008/10/P10008; Bonnet D, 1997, NAT MED, V3, P730, DOI 10.1038/nm0797-730; Collins AT, 2005, CANCER RES, V65, P10946, DOI 10.1158/0008-5472.CAN-05-2018; Craig FE, 2008, BLOOD, V111, P3941, DOI 10.1182/blood-2007-11-120535; Eppert K, 2011, NAT MED, V17, P1086, DOI 10.1038/nm.2415; Finck R, 2013, CYTOM PART A, V83A, P483, DOI 10.1002/cyto.a.22271; Fortunato S, 2010, PHYS REP, V486, P75, DOI 10.1016/j.physrep.2009.11.002; Gibbs KD, 2011, BLOOD, V117, P4226, DOI 10.1182/blood-2010-07-298232; Irish JM, 2004, CELL, V118, P217, DOI 10.1016/j.cell.2004.06.028; Jaatinen T, 2006, STEM CELLS, V24, P631, DOI 10.1634/stemcells.2005-0185; Lu P, 2003, P NATL ACAD SCI USA, V100, P10370, DOI 10.1073/pnas.1832361100; Marusyk A, 2012, NAT REV CANCER, V12, P323, DOI 10.1038/nrc3261; Metzeler KH, 2008, BLOOD, V112, P4193, DOI 10.1182/blood-2008-02-134411; O'Brien CA, 2007, NATURE, V445, P106, DOI 10.1038/nature05372; Patel AP, 2014, SCIENCE, V344, P1396, DOI 10.1126/science.1254257; Pearce DJ, 2006, BLOOD, V107, P1166, DOI 10.1182/blood-2005-06-2325; Pyne S, 2009, P NATL ACAD SCI USA, V106, P8519, DOI 10.1073/pnas.0903028106; Radtke I, 2009, P NATL ACAD SCI USA, V106, P12944, DOI 10.1073/pnas.0903142106; Subramanian A, 2005, P NATL ACAD SCI USA, V102, P15545, DOI 10.1073/pnas.0506580102; Taussig DC, 2008, BLOOD, V112, P568, DOI 10.1182/blood-2007-10-118331; Taussig DC, 2010, BLOOD, V115, P1976, DOI 10.1182/blood-2009-02-206565; Tenen DG, 2003, NAT REV CANCER, V3, P89, DOI 10.1038/nrc989; Toren A, 2005, STEM CELLS, V23, P1142, DOI 10.1634/stemcells.2004-0317; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; Zunder ER, 2015, NAT PROTOC, V10, P316, DOI 10.1038/nprot.2015.020	29	413	414	6	44	CELL PRESS	CAMBRIDGE	50 HAMPSHIRE ST, FLOOR 5, CAMBRIDGE, MA 02139 USA	0092-8674	1097-4172		CELL	Cell	JUL 2	2015	162	1					184	197		10.1016/j.cell.2015.05.047			14	Biochemistry & Molecular Biology; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	CM2WF	WOS:000357542300019	26095251	Green Accepted, Bronze			2020-07-01	
J	Bendall, SC; Davis, KL; Amir, ED; Tadmor, MD; Simonds, EF; Chen, TJ; Shenfeld, DK; Nolan, GP; Pe'er, D				Bendall, Sean C.; Davis, Kara L.; Amir, El-ad David; Tadmor, Michelle D.; Simonds, Erin F.; Chen, Tiffany J.; Shenfeld, Daniel K.; Nolan, Garry P.; Pe'er, Dana			Single-Cell Trajectory Detection Uncovers Progression and Regulatory Coordination in Human B Cell Development	CELL			English	Article							MASS CYTOMETRY; IMMUNOGLOBULIN; LYMPHOCYTES; MURINE; STAT5	Tissue regeneration is an orchestrated progression of cells from an immature state to amature one, conventionally represented as distinctive cell subsets. A continuum of transitional cell states exists between these discrete stages. We combine the depth of single-cell mass cytometry and an algorithm developed to leverage this continuumby aligning single cells of a given lineage onto a unified trajectory that accurately predicts the developmental path de novo. Applied to human B cell lymphopoiesis, the algorithm (termed Wanderlust) constructed trajectories spanning from hematopoietic stem cells through to naive B cells. This trajectory revealed nascent fractions of B cell progenitors and aligned them with developmentally cued regulatory signaling including IL-7/STAT5 and cellular events such as immunoglobulin rearrangement, highlighting checkpoints across which regulatory signals are rewired paralleling changes in cellular state. This study provides a comprehensive analysis of human B lymphopoiesis, laying a foundation to apply this approach to other tissues and "corrupted'' developmental processes including cancer.	[Bendall, Sean C.; Davis, Kara L.; Simonds, Erin F.; Chen, Tiffany J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Bendall, Sean C.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Davis, Kara L.] Stanford Univ, Dept Pediat, Stanford, CA 94305 USA; [Amir, El-ad David; Tadmor, Michelle D.; Shenfeld, Daniel K.; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA; [Chen, Tiffany J.] Stanford Univ, Program Biomed Informat, Stanford, CA 94305 USA; [Chen, Tiffany J.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA[Bendall, Sean C.; Davis, Kara L.; Simonds, Erin F.; Chen, Tiffany J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Bendall, Sean C.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Davis, Kara L.] Stanford Univ, Dept Pediat, Stanford, CA 94305 USA; [Amir, El-ad David; Tadmor, Michelle D.; Shenfeld, Daniel K.; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA; [Chen, Tiffany J.] Stanford Univ, Program Biomed Informat, Stanford, CA 94305 USA; [Chen, Tiffany J.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edu; dpeer@biology.columbia.edugnolan@stanford.edu; dpeer@biology.columbia.edu	Nolan, Garry/AAE-7903-2019	Pe'er, Dana/0000-0002-9259-8817; Simonds, Erin/0000-0002-3497-4861	St. Baldrick's Foundation Scholar Award; Alex's Lemonade Stand Young Investigator Award; Lucile Packard Foundation for Children's Health; NIH-NCATS-CTSAUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Center for Advancing Translational Sciences (NCATS) [UL1 TR001085]; Child Health Research Institute of Stanford University; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [K99GM104148-01, S10 SIG S10RR027582-01, 152175.5041015.0412, DP2-OD002414-01, U54CA121852-01A1]; National Defense Science & Engineering Graduate Fellowship Program; Stanford University Graduate FellowshipStanford University; Google Anita Borg Memorial Scholarship; William Lawrence and Blanche Hughes Foundation; Entertainment Industry Foundation; Northrup-Grumman Corp; Alliance for Lupus Research; Lymphoma Research Foundation; Bill and Melinda Gates FoundationGates Foundation; NSFNational Science Foundation (NSF) [MCB-1149728]; Packard Fellowship for Science and Engineering;  [0158 G KB065];  [1R01CA130826];  [5U54CA143907NIH];  [CIRM: DR1-01477];  [HEALTH.2010.1.2-1];  [HHSF223201210194C - FDA: BAA-12-00118];  [HHSN272200700038C];  [N01-HV-00242];  [NIH 41000411217];  [NIH 5-24927];  [P01 CA034233-22A1];  [PN2EY018228];  [RB2-01592];  [U19 AI057229];  [U54CA149145];  [W81XWH-12-1-0591]	We would like to thank Omer Angel, Antonio de-la-Hera, Astraea Jager, Ulf Klein, Smita Krishnaswamy, Jacob Levine, Eva Sanz, Peter Sims, and Angelica Trejo for their intellectual and technical contributions. K.L.D. is supported by a St. Baldrick's Foundation Scholar Award, Alex's Lemonade Stand Young Investigator Award and the Lucile Packard Foundation for Children's Health, Stanford NIH-NCATS-CTSA UL1 TR001085 and Child Health Research Institute of Stanford University. S.C.B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09) and the NIH K99GM104148-01. E.D.A. is a Howard Hughes Medical Institute International Student Research Fellow. T.J.C. is supported by National Defense Science & Engineering Graduate Fellowship Program, Stanford University Graduate Fellowship, and the Google Anita Borg Memorial Scholarship. This work was supported by 0158 G KB065; 1R01CA130826; 5U54CA143907NIH; CIRM: DR1-01477; HEALTH.2010.1.2-1; HHSF223201210194C - FDA: BAA-12-00118; HHSN272200700038C; N01-HV-00242; NIH 41000411217; NIH 5-24927; P01 CA034233-22A1; PN2EY018228; RB2-01592; U19 AI057229; U54CA149145; W81XWH-12-1-0591 OCRP-TIA NWC; NIH S10 SIG S10RR027582-01, William Lawrence and Blanche Hughes Foundation, Entertainment Industry Foundation, Northrup-Grumman Corp, Alliance for Lupus Research, Lymphoma Research Foundation, Bill and Melinda Gates Foundation, NIH 152175.5041015.0412 and the Rachford and Carlota A. Harris Endowment to G.P.N. This work was supported by NSF MCB-1149728, NIH DP2-OD002414-01, NIH U54CA121852-01A1 to D.P. D.P. holds a Packard Fellowship for Science and Engineering. G.P.N. has personal financial interest in and S.C.B. is a paid consultant for the company DVS Sciences, the manufacturers that produced some of the reagents and instrumentation used in this manuscript.	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J	Newell, EW; Sigal, N; Bendall, SC; Nolan, GP; Davis, MM				Newell, Evan W.; Sigal, Natalia; Bendall, Sean C.; Nolan, Garry P.; Davis, Mark M.			Cytometry by Time-of-Flight Shows Combinatorial Cytokine Expression and Virus-Specific Cell Niches within a Continuum of CD8(+) T Cell Phenotypes	IMMUNITY			English	Article							CLASS-I LIGANDS; EFFECTOR FUNCTIONS; PEPTIDE COMPLEXES; MHC TETRAMERS; MEMORY; ANTIGEN; LYMPHOCYTES; SUBSETS; RESPONSES; IMMUNE	Cytotoxic CD8(+) T lymphocytes directly kill infected or aberrant cells and secrete proinflammatory cytokines. By using metal-labeled probes and mass spectrometric analysis (cytometry by time-of-flight, or CyTOF) of human CD8(+) T cells, we analyzed the expression of many more proteins than previously possible with fluorescent labels, including surface markers, cytokines, and antigen specificity with modified peptide-MHC tetramers. With 3-dimensional principal component analysis (3D-PCA) to display phenotypic diversity, we observed a relatively uniform pattern of variation in all subjects tested, highlighting the interrelatedness of previously described subsets and the continuous nature of CD8(+) T cell differentiation. These data also showed much greater complexity in the CD8(+) T cell compartment than previously appreciated, including a nearly combinatorial pattern of cytokine expression, with distinct niches occupied by virus-specific cells. This large degree of functional diversity even between cells with the same specificity gives CD8(+) T cells a remarkable degree of flexibility in responding to pathogens.	[Newell, Evan W.; Bendall, Sean C.; Nolan, Garry P.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Sigal, Natalia; Nolan, Garry P.; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Stanford, CA 94305 USA; [Davis, Mark M.] Stanford Univ, Howard Hughes Med Inst, Stanford, CA 94305 USA[Newell, Evan W.; Bendall, Sean C.; Nolan, Garry P.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Sigal, Natalia; Nolan, Garry P.; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Stanford, CA 94305 USA; [Davis, Mark M.] Stanford Univ, Howard Hughes Med Inst, Stanford, CA 94305 USA	Davis, MM (reprint author), Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA.	mmdavis@stanford.edummdavis@stanford.edu	Newell, Evan W/F-9711-2012; Newell, Evan/AAE-9470-2020; Nolan, Garry/AAE-7903-2019	Newell, Evan W/0000-0002-2889-243X; Newell, Evan/0000-0002-2889-243X; 	NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19-AI057229, U19-AI090019]; Bill and Melinda Gates FoundationGates Foundation; Howard Hughes Medical InstituteHoward Hughes Medical Institute; American Cancer SocietyAmerican Cancer Society; Damon Runyon Postdoctoral Fellowship [DRG-2017-09]	The authors would like to thank D. Parks and W. Moore for helpful discussions and help with FlowJo and FCS files, P. Lund for helpful discussions and a T cell stimulation protocol, M. Leipold for help with CyTOF machine maintenance, E. Zunder, B. Bodenmiller, and E. Simonds for general help with CyTOF usage, and A. Han, B. Kidd, W. O'Gorman, O. Goldberger, and Y.-h. Chien for helpful discussions. This work was supported by NIH grants U19-AI057229 and U19-AI090019, Bill and Melinda Gates Foundation Grand Challenges Exploration phase land II grants, and The Howard Hughes Medical Institute. E.W.N. is supported by The American Cancer Society's Steven Stanley and Edward Albert Bielfelt Post-Doctoral Fellowship. S.C.B. is supported by a Damon Runyon Postdoctoral Fellowship (DRG-2017-09). G.P.N. is supported by an endowed chair from Rachtford and Carlota A. Harris. G.P.N. is a paid consultant for Becton Dickinson Biosciences, a member of the Board of Directors and consultant for DVS Sciences, and has equity holdings in DVS Biosciences.	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J	Bendall, SC; Nolan, GP; Roederer, M; Chattopadhyay, PK				Bendall, Sean C.; Nolan, Garry P.; Roederer, Mario; Chattopadhyay, Pratip K.			A deep profiler's guide to cytometry	TRENDS IN IMMUNOLOGY			English	Review						fluorescence; inductively coupled plasma mass spectrometry; single cell analysis; immunophenotyping; data analysis	ELEMENT-TAGGED IMMUNOASSAY; FLOW-CYTOMETRY; MASS CYTOMETRY; CYTOKINE EXPRESSION; MULTIPLEX BIOASSAY; T-CELLS; POLYMER; IDENTIFICATION; PHYCOERYTHRIN; LYMPHOCYTES	In recent years, advances in technology have provided us with tools to quantify the expression of multiple genes in individual cells. The ability to measure simultaneously multiple genes in the same cell is necessary to resolve the great diversity of cell subsets, as well as to define their function in the host. Fluorescence-based flow cytometry is the benchmark for this; with it, we can quantify 18 proteins per cell, at >10 000 cells/s. Mass cytometry is a new technology that promises to extend these capabilities significantly. Immunophenotyping by mass spectrometry provides the ability to measure >36 proteins at a rate of 1000 cells/s. We review these cytometric technologies, capable of high-content, high-throughput single-cell assays.	[Roederer, Mario; Chattopadhyay, Pratip K.] NIAID, ImmunoTechnol Sect, Vaccine Res Ctr, NIH, Bethesda, MD 20892 USA; [Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Baxter Lab Stem Cell Biol, Dept Microbiol & Immunol, Stanford, CA 94305 USA[Roederer, Mario; Chattopadhyay, Pratip K.] NIAID, ImmunoTechnol Sect, Vaccine Res Ctr, NIH, Bethesda, MD 20892 USA; [Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Baxter Lab Stem Cell Biol, Dept Microbiol & Immunol, Stanford, CA 94305 USA	Roederer, M (reprint author), NIAID, ImmunoTechnol Sect, Vaccine Res Ctr, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.	Roederer@nih.govRoederer@nih.gov	Nolan, Garry/AAE-7903-2019	Chattopadhyay, Pratip/0000-0002-5457-9666	Intramural Research Program of the NIAID, NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID); Collaboration for AIDS Vaccine Discovery (CAVD), from the Bill and Melinda Gates Foundation [OPP1032325]; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; Rachford and Carlota A. Harris Endowed Professorship; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; Bill and Melinda Gates FoundationGates Foundation [GF12141-137101];  [U19 AI057229];  [P01 CA034233];  [HHSN272200700038C];  [1R01CA130826];  [CIRM DR1-01477];  [RB2-01592];  [NCI RFA CA 09-011];  [NHLBI-HV-10-05(2)]	M.R. and P.K.C. are supported by the Intramural Research Program of the NIAID, NIH, and by the Collaboration for AIDS Vaccine Discovery (CAVD), Grant #OPP1032325, from the Bill and Melinda Gates Foundation. S.C.B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09). G.P.N. is supported by the Rachford and Carlota A. Harris Endowed Professorship and grants from U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, CIRM DR1-01477 and RB2-01592, NCI RFA CA 09-011, NHLBI-HV-10-05(2), European Commission HEALTH.2010.1.2-1, and the Bill and Melinda Gates Foundation (GF12141-137101).	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JUL	2012	33	7					323	332		10.1016/j.it.2012.02.010			10	Immunology	Immunology	977EP	WOS:000306639500001	22476049	Green Accepted			2020-07-01	
J	Spitzer, MH; Nolan, GP				Spitzer, Matthew H.; Nolan, Garry P.			Mass Cytometry: Single Cells, Many Features	CELL			English	Article							FLOW-CYTOMETRY; IMMUNE-SYSTEM; IDENTIFICATION; EXPRESSION; PROGRESSION; DIVERSITY; RESPONSES	Technology development in biological research often aims to either increase the number of cellular features that can be surveyed simultaneously or enhance the resolution at which such observations are possible. For decades, flow cytometry has balanced these goals to fill a critical need by enabling the measurement of multiple features in single cells, commonly to examine complex or hierarchical cellular systems. Recently, a format for flow cytometry has been developed that leverages the precision of mass spectrometry. This fusion of the two technologies, termed mass cytometry, provides measurement of over 40 simultaneous cellular parameters at single-cell resolution, significantly augmenting the ability of cytometry to evaluate complex cellular systems and processes. In this Primer, we review the current state of mass cytometry, providing an overview of the instrumentation, its present capabilities, and methods of data analysis, as well as thoughts on future developments and applications.	[Spitzer, Matthew H.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Nolan, Garry P.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA[Spitzer, Matthew H.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Nolan, Garry P.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA	Spitzer, MH; Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.; Spitzer, MH; Nolan, GP (reprint author), Stanford Univ, Program Immunol, Stanford, CA 94305 USA.; Spitzer, MH (reprint author), Stanford Univ, Dept Pathol, Stanford, CA 94305 USA.; Spitzer, MH (reprint author), Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA.	matthew.spitzer@ucsf.edu; gnolan@stanford.edumatthew.spitzer@ucsf.edu; gnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Spitzer, Matthew/0000-0002-5291-3819	company Fluidigm	G.P.N. has a personal financial interest in the company Fluidigm, the manufacturer of the mass cytometer referenced in this study.	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J	Bodenmiller, B; Zunder, ER; Finck, R; Chen, TJ; Savig, ES; Bruggner, RV; Simonds, EF; Bendall, SC; Sachs, K; Krutzik, PO; Nolan, GP				Bodenmiller, Bernd; Zunder, Eli R.; Finck, Rachel; Chen, Tiffany J.; Savig, Erica S.; Bruggner, Robert V.; Simonds, Erin F.; Bendall, Sean C.; Sachs, Karen; Krutzik, Peter O.; Nolan, Garry P.			Multiplexed mass cytometry profiling of cellular states perturbed by small-molecule regulators	NATURE BIOTECHNOLOGY			English	Article							KINASE INHIBITOR SELECTIVITY; POLYCHROMATIC FLOW-CYTOMETRY; FUNCTIONAL GENOMICS; DRUG DISCOVERY; IMMUNE-SYSTEM; CANCER; CELLS; PROTEOMICS; RESPONSES; POPULATIONS	Mass cytometry facilitates high-dimensional, quantitative analysis of the effects of bioactive molecules on human samples at single-cell resolution, but instruments process only one sample at a time. Here we describe mass-tag cellular barcoding (MCB), which increases mass cytometry throughput by using n metal ion tags to multiplex up to 2(n) samples. We used seven tags to multiplex an entire 96-well plate, and applied MCB to characterize human peripheral blood mononuclear cell (PBMC) signaling dynamics and cell-to-cell communication, signaling variability between PBMCs from eight human donors, and the effects of 27 inhibitors on this system. For each inhibitor, we measured 14 phosphorylation sites in 14 PBMC types at 96 conditions, resulting in 18,816 quantified phosphorylation levels from each multiplexed sample. This high-dimensional, systems-level inquiry allowed analysis across cell-type and signaling space, reclassified inhibitors and revealed off-target effects. High-content, high-throughput screening with MCB should be useful for drug discovery, preclinical testing and mechanistic investigation of human disease.	[Bodenmiller, Bernd; Zunder, Eli R.; Finck, Rachel; Chen, Tiffany J.; Savig, Erica S.; Bruggner, Robert V.; Simonds, Erin F.; Bendall, Sean C.; Sachs, Karen; Krutzik, Peter O.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Chen, Tiffany J.; Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA; [Chen, Tiffany J.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Savig, Erica S.] Stanford Univ, Canc Biol Program, Stanford, CA 94305 USA[Bodenmiller, Bernd; Zunder, Eli R.; Finck, Rachel; Chen, Tiffany J.; Savig, Erica S.; Bruggner, Robert V.; Simonds, Erin F.; Bendall, Sean C.; Sachs, Karen; Krutzik, Peter O.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Chen, Tiffany J.; Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA; [Chen, Tiffany J.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Savig, Erica S.] Stanford Univ, Canc Biol Program, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Simonds, Erin/0000-0002-3497-4861	Swiss National Science Foundation (SNF)Swiss National Science Foundation (SNSF); European Molecular Biology Organization (EMBO)European Molecular Biology Organization (EMBO); Marie Curie IOFEuropean Union (EU); National Institute of General Medical SciencesUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of General Medical Sciences (NIGMS) [F32GM093508]; Department of Defense (DoD) through the National Defense Science & Engineering Graduate Fellowship (NDSEG) Program; Stanford Graduate Fellowship in Science and Engineering; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; Rachford and Carlota A. Harris Endowed Professorship; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; Bill and Melinda Gates FoundationGates Foundation [GF12141-137101];  [U19 AI057229];  [P01 CA034233];  [HHSN272200700038C];  [1R01CA130826];  [CIRM DR1-01477];  [RB2-01592];  [NCI RFA CA 09-011];  [NHLBI-HV-10-05(2)]	We would like to thank A. Trejo, M. Clutter, K. Gibbs and G. Behbahani for their experimental support and discussions, and D. Pe'er and El-ad D. Amir for their feedback on data analysis. B. B. was supported by fellowships of the Swiss National Science Foundation (SNF), the European Molecular Biology Organization (EMBO), and the Marie Curie IOF. E.R.Z. is supported by a fellowship from National Institute of General Medical Sciences (F32GM093508). T.J.C. is supported by the Department of Defense (DoD) through the National Defense Science & Engineering Graduate Fellowship (NDSEG) Program, and the Stanford Graduate Fellowship in Science and Engineering. S. C. B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09). G.P.N. is supported by the Rachford and Carlota A. Harris Endowed Professorship and grants from U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, CIRM DR1-01477 and RB2-01592, NCI RFA CA 09-011, NHLBI-HV-10-05(2), European Commission HEALTH.2010.1.2-1, and the Bill and Melinda Gates Foundation (GF12141-137101).	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Biotechnol.	SEP	2012	30	9					858	U89		10.1038/nbt.2317			12	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	004UD	WOS:000308705700020	22902532	Green Accepted			2020-07-01	
J	Wei, SC; Levine, JH; Cogdill, AP; Zhao, Y; Anang, NAAS; Andrews, MC; Sharma, P; Wang, J; Wargo, JA; Pe'er, D; Allison, JP				Wei, Spencer C.; Levine, Jacob H.; Cogdill, Alexandria P.; Zhao, Yang; Anang, Nana-Ama A. S.; Andrews, Miles C.; Sharma, Padmanee; Wang, Jing; Wargo, Jennifer A.; Pe'er, Dana; Allison, James P.			Distinct Cellular Mechanisms Underlie Anti-CTLA-4 and Anti-PD-1 Checkpoint Blockade	CELL			English	Article							REGULATORY T-CELLS; MASS CYTOMETRY; CANCER-IMMUNOTHERAPY; CTLA-4 BLOCKADE; COMBINATION IMMUNOTHERAPY; METASTATIC MELANOMA; PROGRAMMED DEATH-1; SOLID TUMORS; THERAPY; IMMUNE	Immune-checkpoint blockade is able to achieve durable responses in a subset of patients; however, we lack a satisfying comprehension of the underlying mechanisms of anti-CTLA-4- and anti-PD-1-induced tumor rejection. To address these issues, we utilized mass cytometry to comprehensively profile the effects of checkpoint blockade on tumor immune infiltrates in human melanoma and murine tumor models. These analyses reveal a spectrum of tumor-infiltrating T cell populations that are highly similar between tumor models and indicate that checkpoint blockade targets only specific subsets of tumor-infiltrating T cell populations. Anti-PD-1 predominantly induces the expansion of specific tumor-infiltrating exhausted-like CD8 T cell subsets. In contrast, anti-CTLA-4 induces the expansion of an ICOS+ Th1-like CD4 effector population in addition to engaging specific subsets of exhaustedlike CD8 T cells. Thus, our findings indicate that anti-CTLA-4 and anti-PD-1 checkpoint-blockade-induced immune responses are driven by distinct cellular mechanisms.	[Wei, Spencer C.; Cogdill, Alexandria P.; Anang, Nana-Ama A. S.; Allison, James P.] Univ Texas MD Anderson Canc Ctr, Dept Immunol, Houston, TX 77030 USA; [Levine, Jacob H.; Pe'er, Dana] Sloan Kettering Inst, Computat & Syst Biol Program, New York, NY 10065 USA; [Cogdill, Alexandria P.; Andrews, Miles C.; Wargo, Jennifer A.] Univ Texas MD Anderson Canc Ctr, Dept Surg Oncol, Houston, TX 77030 USA; [Zhao, Yang; Wang, Jing] Univ Texas MD Anderson Canc Ctr, Dept Bioinformat & Computat Biol, Houston, TX 77030 USA; [Sharma, Padmanee] Univ Texas MD Anderson Canc Ctr, Dept Genitourinary Med Oncol, Houston, TX 77030 USA; [Sharma, Padmanee; Wargo, Jennifer A.; Allison, James P.] Univ Texas MD Anderson Canc Ctr, Parker Inst Canc Immunotherapy, Houston, TX 77030 USA; [Wargo, Jennifer A.] Univ Texas MD Anderson Canc Ctr, Dept Genom Med, Houston, TX 77030 USA[Wei, Spencer C.; Cogdill, Alexandria P.; Anang, Nana-Ama A. S.; Allison, James P.] Univ Texas MD Anderson Canc Ctr, Dept Immunol, Houston, TX 77030 USA; [Levine, Jacob H.; Pe'er, Dana] Sloan Kettering Inst, Computat & Syst Biol Program, New York, NY 10065 USA; [Cogdill, Alexandria P.; Andrews, Miles C.; Wargo, Jennifer A.] Univ Texas MD Anderson Canc Ctr, Dept Surg Oncol, Houston, TX 77030 USA; [Zhao, Yang; Wang, Jing] Univ Texas MD Anderson Canc Ctr, Dept Bioinformat & Computat Biol, Houston, TX 77030 USA; [Sharma, Padmanee] Univ Texas MD Anderson Canc Ctr, Dept Genitourinary Med Oncol, Houston, TX 77030 USA; [Sharma, Padmanee; Wargo, Jennifer A.; Allison, James P.] Univ Texas MD Anderson Canc Ctr, Parker Inst Canc Immunotherapy, Houston, TX 77030 USA; [Wargo, Jennifer A.] Univ Texas MD Anderson Canc Ctr, Dept Genom Med, Houston, TX 77030 USA	Wei, SC; Allison, JP (reprint author), Univ Texas MD Anderson Canc Ctr, Dept Immunol, Houston, TX 77030 USA.; Allison, JP (reprint author), Univ Texas MD Anderson Canc Ctr, Parker Inst Canc Immunotherapy, Houston, TX 77030 USA.	scwei@mdanderson.org; jallison@mdanderson.orgscwei@mdanderson.org; jallison@mdanderson.org		Cogdill, Alexandria/0000-0001-8917-9462; Pe'er, Dana/0000-0002-9259-8817; Anang, Nana-Ama/0000-0002-8495-2978; Allison, James/0000-0001-8980-5697; Wei, Spencer/0000-0001-9673-9683; Andrews, Miles/0000-0003-1231-8641	NCI Cancer Center Support Grant (CCSG) [P30CA016672]; Cancer Prevention and Research Institute of Texas (CPRIT) [R1203]; MDACC CCSG Bioinformatics Shared Resource [P30CA016672]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DP1-HD084071, R01CA164729, R01CA163793]; MSK Cancer Center Support Grant/Core Grant [P30CA008748]	We thank Duncan Mak for expert advice and technical assistance with mass cytometry analysis. Mass cytometry and cell sorting were performed at the MDACC Flow Cytometry and Cellular Imaging Core Facility, which is in part funded by NCI Cancer Center Support Grant (CCSG) P30CA016672. Sequencing was performed at the MDACC Sequencing and Microarray Facility, which is also in part funded by P30CA016672(SMF). This work was supported by a grant from Cancer Prevention and Research Institute of Texas (CPRIT) to J.P.A. (R1203), the MDACC CCSG Bioinformatics Shared Resource (P30CA016672 39), NIH grants to D.P. (DP1-HD084071 and R01CA164729), and a MSK Cancer Center Support Grant/Core Grant to D.P. (P30CA008748). S.C.W. is an MDACC Odyssey postdoctoral fellow. J.P.A. and P.S. are co-directors, and J.A.W. is a member of the Parker Institute for Cancer Immunotherapy. P.S. has consulted for AstraZeneca, Amgen, GlaxoSmithKline (GSK), and Bristol-Myers Squibb (BMS). P.S. serves on the scientific advisory boards for Jounce, Neon, Constellation, and Kite. P.S. is a co-founder of Jounce Therapeutics. P.S. received funding on NIH R01CA163793. M.C.A. has attended speakers' bureau with travel support from BMS and MSD. J.A.W. has received honoraria from speakers' bureau of Dava Oncology, BMS, and Illumina and is an advisory board member for GSK, Novartis, and Roche/Genentech. J.P.A. is a co-founder of Jounce and Neon Therapeutics, is an advisory board member for Jounce, Neon, Amgen, and Kite Pharmaceuticals, and has received royalties from intellectual property licensed to BMS and Merck.	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J	Guilliams, M; Dutertre, CA; Scott, CL; McGovern, N; Sichien, D; Chakarov, S; Van Gassen, S; Chen, JM; Poidinger, M; De Prijck, S; Tavernier, SJ; Low, I; Irac, SE; Mattar, CN; Sumatoh, HR; Low, GHL; Chung, TJK; Chan, DKH; Tan, KK; Hon, TLK; Fossum, E; Bogen, B; Choolani, M; Chan, JKY; Larbi, A; Luche, H; Henri, S; Saeys, Y; Newell, EW; Lambrecht, BN; Malissen, B; Ginhoux, F				Guilliams, Martin; Dutertre, Charles-Antoine; Scott, Charlotte L.; McGovern, Naomi; Sichien, Dorine; Chakarov, Svetoslav; Van Gassen, Sofie; Chen, Jinmiao; Poidinger, Michael; De Prijck, Sofie; Tavernier, Simon J.; Low, Ivy; Irac, Sergio Erdal; Mattar, Citra Nurfarah; Sumatoh, Hermi Rizal; Low, Gillian Hui Ling; Chung, Tam John Kit; Chan, Dedrick Kok Hong; Tan, Ker Kan; Hon, Tony Lim Kiat; Fossum, Even; Bogen, Bjame; Choolani, Mahesh; Chan, Jerry Kok Yen; Larbi, Anis; Luche, Herve; Henri, Sandrine; Saeys, Yvan; Newell, Evan William; Lambrecht, Bart N.; Malissen, Bernard; Ginhoux, Florent			Unsupervised High-Dimensional Analysis Aligns Dendritic Cells across Tissues and Species	IMMUNITY			English	Article							CLONOGENIC PROGENITOR; SIGNALING CONTROLS; STEADY-STATE; LYMPH-NODES; BONE-MARROW; MACROPHAGES; EXPRESSION; RESPONSES; BLOOD; HOMEOSTASIS	Dendritic cells (DCs) are professional antigen-presenting cells that hold great therapeutic potential. Multiple DC subsets have been described, and it remains challenging to align them across tissues and species to analyze their function in the absence of macrophage contamination. Here, we provide and validate a universal toolbox for the automated identification of DCs through unsupervised analysis of conventional flow cytometry and mass cytometry data obtained from multiple mouse, macaque, and human tissues. The use of a minimal set of lineage-imprinted markers was sufficient to subdivide DCs into conventional type 1 (cDC1s), conventional type 2 (cDC2s), and plasmacytoid DCs (pDCs) across tissues and species. This way, a large number of additional markers can still be used to further characterize the heterogeneity of DCs across tissues and during inflammation. This framework represents the way forward to a universal, high-throughput, and standardized analysis of DC populations from mutant mice and human patients.	[Guilliams, Martin; Scott, Charlotte L.; Sichien, Dorine; De Prijck, Sofie; Tavernier, Simon J.; Lambrecht, Bart N.] VIB Inflammat Res Ctr, Unit Immunoregulat & Mucosal Immunol, B-9052 Ghent, Belgium; [Guilliams, Martin; Scott, Charlotte L.; Sichien, Dorine] Univ Ghent, Dept Biomed Mol Biol, B-9000 Ghent, Belgium; [Guilliams, Martin; Luche, Herve; Henri, Sandrine; Malissen, Bernard] Aix Marseille Univ, CNRS, INSERM, Ctr Immunol Marseille Luminy, F-13288 Marseille, France; [Dutertre, Charles-Antoine; McGovern, Naomi; Chakarov, Svetoslav; Chen, Jinmiao; Poidinger, Michael; Low, Ivy; Sumatoh, Hermi Rizal; Low, Gillian Hui Ling; Chan, Jerry Kok Yen; Larbi, Anis; Newell, Evan William; Ginhoux, Florent] ASTAR, Singapore Immunol Network SIgN, BIOPOLIS, 8A Biomed Grove,IMMUNOS Bldg 3-4, Singapore 138648, Singapore; [Dutertre, Charles-Antoine; Irac, Sergio Erdal] Duke NUS Med Sch, Program Emerging Infect Dis, 8 Coll Rd, Singapore 169857, Singapore; [Van Gassen, Sofie] Univ Ghent, Dept Informat Technol, iMinds, B-9000 Ghent, Belgium; [Van Gassen, Sofie; Saeys, Yvan] VIB Inflammat Res Ctr, Data Min & Modeling Biomed, B-9052 Ghent, Belgium; [De Prijck, Sofie; Tavernier, Simon J.; Saeys, Yvan; Lambrecht, Bart N.] Univ Ghent, Dept Internal Med, B-9000 Ghent, Belgium; [Mattar, Citra Nurfarah; Choolani, Mahesh; Chan, Jerry Kok Yen] Natl Univ Singapore, Yong Loo Lin Sch Med, Expt Fetal Med Grp, Singapore 119077, Singapore; [Chung, Tam John Kit; Chan, Dedrick Kok Hong; Tan, Ker Kan] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Surg, Singapore 119077, Singapore; [Hon, Tony Lim Kiat] Natl Univ Singapore, Dept Pathol, Singapore 119077, Singapore; [Fossum, Even; Bogen, Bjame] Univ Oslo, Oslo Univ Hosp, KG Jebsen Ctr Influenza Vaccine Res, N-0027 Oslo, Norway; [Bogen, Bjame] Univ Oslo, Oslo Univ Hosp, Rikshosp, Inst Immunol,Ctr Immune Regulat, N-0424 Oslo, Norway; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Div Obstet & Gynaecol, Singapore 229899, Singapore; [Chan, Jerry Kok Yen] Duke NUS Grad Med Sch, Canc & Stem Cell Biol Program, Singapore 119077, Singapore; [Luche, Herve; Malissen, Bernard] Aix Marseille Univ, CNRS, Ctr Immunophenom, INSERM, F-13288 Marseille, France; [Lambrecht, Bart N.] Erasmus MC, Dept Pulm Med, Dr Molewaterpl 50, NL-3015 GE Rotterdam, Netherlands[Guilliams, Martin; Scott, Charlotte L.; Sichien, Dorine; De Prijck, Sofie; Tavernier, Simon J.; Lambrecht, Bart N.] VIB Inflammat Res Ctr, Unit Immunoregulat & Mucosal Immunol, B-9052 Ghent, Belgium; [Guilliams, Martin; Scott, Charlotte L.; Sichien, Dorine] Univ Ghent, Dept Biomed Mol Biol, B-9000 Ghent, Belgium; [Guilliams, Martin; Luche, Herve; Henri, Sandrine; Malissen, Bernard] Aix Marseille Univ, CNRS, INSERM, Ctr Immunol Marseille Luminy, F-13288 Marseille, France; [Dutertre, Charles-Antoine; McGovern, Naomi; Chakarov, Svetoslav; Chen, Jinmiao; Poidinger, Michael; Low, Ivy; Sumatoh, Hermi Rizal; Low, Gillian Hui Ling; Chan, Jerry Kok Yen; Larbi, Anis; Newell, Evan William; Ginhoux, Florent] ASTAR, Singapore Immunol Network SIgN, BIOPOLIS, 8A Biomed Grove,IMMUNOS Bldg 3-4, Singapore 138648, Singapore; [Dutertre, Charles-Antoine; Irac, Sergio Erdal] Duke NUS Med Sch, Program Emerging Infect Dis, 8 Coll Rd, Singapore 169857, Singapore; [Van Gassen, Sofie] Univ Ghent, Dept Informat Technol, iMinds, B-9000 Ghent, Belgium; [Van Gassen, Sofie; Saeys, Yvan] VIB Inflammat Res Ctr, Data Min & Modeling Biomed, B-9052 Ghent, Belgium; [De Prijck, Sofie; Tavernier, Simon J.; Saeys, Yvan; Lambrecht, Bart N.] Univ Ghent, Dept Internal Med, B-9000 Ghent, Belgium; [Mattar, Citra Nurfarah; Choolani, Mahesh; Chan, Jerry Kok Yen] Natl Univ Singapore, Yong Loo Lin Sch Med, Expt Fetal Med Grp, Singapore 119077, Singapore; [Chung, Tam John Kit; Chan, Dedrick Kok Hong; Tan, Ker Kan] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Surg, Singapore 119077, Singapore; [Hon, Tony Lim Kiat] Natl Univ Singapore, Dept Pathol, Singapore 119077, Singapore; [Fossum, Even; Bogen, Bjame] Univ Oslo, Oslo Univ Hosp, KG Jebsen Ctr Influenza Vaccine Res, N-0027 Oslo, Norway; [Bogen, Bjame] Univ Oslo, Oslo Univ Hosp, Rikshosp, Inst Immunol,Ctr Immune Regulat, N-0424 Oslo, Norway; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Div Obstet & Gynaecol, Singapore 229899, Singapore; [Chan, Jerry Kok Yen] Duke NUS Grad Med Sch, Canc & Stem Cell Biol Program, Singapore 119077, Singapore; [Luche, Herve; Malissen, Bernard] Aix Marseille Univ, CNRS, Ctr Immunophenom, INSERM, F-13288 Marseille, France; [Lambrecht, Bart N.] Erasmus MC, Dept Pulm Med, Dr Molewaterpl 50, NL-3015 GE Rotterdam, Netherlands	Guilliams, M; Lambrecht, BN (reprint author), VIB Inflammat Res Ctr, Unit Immunoregulat & Mucosal Immunol, B-9052 Ghent, Belgium.; Guilliams, M (reprint author), Univ Ghent, Dept Biomed Mol Biol, B-9000 Ghent, Belgium.; Guilliams, M; Malissen, B (reprint author), Aix Marseille Univ, CNRS, INSERM, Ctr Immunol Marseille Luminy, F-13288 Marseille, France.; Ginhoux, F (reprint author), ASTAR, Singapore Immunol Network SIgN, BIOPOLIS, 8A Biomed Grove,IMMUNOS Bldg 3-4, Singapore 138648, Singapore.; Lambrecht, BN (reprint author), Univ Ghent, Dept Internal Med, B-9000 Ghent, Belgium.; Malissen, B (reprint author), Aix Marseille Univ, CNRS, Ctr Immunophenom, INSERM, F-13288 Marseille, France.; Lambrecht, BN (reprint author), Erasmus MC, Dept Pulm Med, Dr Molewaterpl 50, NL-3015 GE Rotterdam, Netherlands.	martin.guilliams@irc.vib-ugent.be; bart.lambrecht@irc.vib-ugent.be; bernardm@ciml.univ-mrs.fr; florent_ginhoux@immunol.a-star.edu.sgmartin.guilliams@irc.vib-ugent.be; bart.lambrecht@irc.vib-ugent.be; bernardm@ciml.univ-mrs.fr; florent_ginhoux@immunol.a-star.edu.sg	Newell, Evan/AAE-9470-2020; Tavernier, Simon/L-9949-2019; Irac, Sergio Erdal/G-6690-2014; Tan, Ker Kan/K-2506-2017; Fossum, Even/W-9136-2019; Lambrecht, Bart N/K-2484-2014; Saeys, Yvan/C-1311-2009; Henri, Sandrine/O-3045-2016; Newell, Evan W/F-9711-2012; Tam, John/B-6048-2012; Chakarov, Svetoslav/AAC-9283-2019; Guilliams, Martin MG/N-7803-2013; Malissen, Bernard/AAK-4659-2020; Scott, Charlotte/K-3563-2014	Newell, Evan/0000-0002-2889-243X; Tavernier, Simon/0000-0003-2514-5655; Irac, Sergio Erdal/0000-0001-5622-1409; Tan, Ker Kan/0000-0002-0410-0450; Fossum, Even/0000-0001-7064-0327; Lambrecht, Bart N/0000-0003-4376-6834; Saeys, Yvan/0000-0002-0415-1506; Newell, Evan W/0000-0002-2889-243X; Chakarov, Svetoslav/0000-0002-0665-5417; Guilliams, Martin MG/0000-0003-3525-7570; Malissen, Bernard/0000-0003-1340-9342; Van Gassen, Sofie/0000-0002-7119-5330; Scott, Charlotte/0000-0003-4914-6580; Dutertre, Charles-Antoine/0000-0001-7950-3652; Poidinger, Michael/0000-0002-1047-2277; McGovern, Naomi/0000-0001-5200-2698; HENRI, Sandrine/0000-0002-8980-9193	Singapore Immunology NetworkAgency for Science Technology & Research (ASTAR); National Research Foundation Singapore under its cooperative basic research grant new investigator grant [NMRC/BNIG/2026/2014]; Marie Curie Intra-European Fellowship (IEF) as part of Horizon; Marie Curie Reintegration grantEuropean Union (EU); Odysseus grant; FWO grants of the Flemish Government; CNRSCentre National de la Recherche Scientifique (CNRS); INSERMInstitut National de la Sante et de la Recherche Medicale (Inserm); PHENOMIN; European Research Council (FP7) [322465]; Flanders Agency for Innovation by Science and Technology (IWT)Institute for the Promotion of Innovation by Science and Technology in Flanders (IWT)	We thank Gert Van Isterdael and Charlene Foong Shu Fen from the Flow Cytometry Core Platform of the IRC and SingHealth, respectively. This work was supported by core grants of the Singapore Immunology Network to F.G. and E.W.N. and by the National Research Foundation Singapore under its cooperative basic research grant new investigator grant (NMRC/BNIG/2026/2014) and administered by the Singapore Ministry of Health's National Medical Research Council to C.-A.D. C.L.S. is supported by a Marie Curie Intra-European Fellowship (IEF) as part of Horizon 2020. M.G. is supported by a Marie Curie Reintegration grant, an Odysseus grant, and FWO grants of the Flemish Government. B.M. is supported by the CNRS, INSERM, PHENOMIN, and European Research Council (FP7/2007-2013 grant no 322465). S.V.G. is funded by the Flanders Agency for Innovation by Science and Technology (IWT).	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J	Finck, R; Simonds, EF; Jager, A; Krishnaswamy, S; Sachs, K; Fantl, W; Pe'er, D; Nolan, GP; Bendall, SC				Finck, Rachel; Simonds, Erin F.; Jager, Astraea; Krishnaswamy, Smita; Sachs, Karen; Fantl, Wendy; Pe'er, Dana; Nolan, Garry P.; Bendall, Sean C.			Normalization of mass cytometry data with bead standards	CYTOMETRY PART A			English	Article						mass cytometry; CyTOF; normalization; blood; PBMC; flow cytometry; phenotype; internal standards		Mass cytometry uses atomic mass spectrometry combined with isotopically pure reporter elements to currently measure as many as 40 parameters per single cell. As with any quantitative technology, there is a fundamental need for quality assurance and normalization protocols. In the case of mass cytometry, the signal variation over time due to changes in instrument performance combined with intervals between scheduled maintenance must be accounted for and then normalized. Here, samples were mixed with polystyrene beads embedded with metal lanthanides, allowing monitoring of mass cytometry instrument performance over multiple days of data acquisition. The protocol described here includes simultaneous measurements of beads and cells on the mass cytometer, subsequent extraction of the bead-based signature, and the application of an algorithm enabling correction of both short- and long-term signal fluctuations. The variation in the intensity of the beads that remains after normalization may also be used to determine data quality. Application of the algorithm to a one-month longitudinal analysis of a human peripheral blood sample reduced the range of median signal fluctuation from 4.9-fold to 1.3-fold. (c) 2013 International Society for Advancement of Cytometry	[Finck, Rachel; Simonds, Erin F.; Jager, Astraea; Sachs, Karen; Fantl, Wendy; Nolan, Garry P.; Bendall, Sean C.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA[Finck, Rachel; Simonds, Erin F.; Jager, Astraea; Sachs, Karen; Fantl, Wendy; Nolan, Garry P.; Bendall, Sean C.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA	Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edu; bendall@stanford.edugnolan@stanford.edu; bendall@stanford.edu	Nolan, Garry/AAE-7903-2019; Krishnaswamy, Smita/D-8089-2016	Krishnaswamy, Smita/0000-0001-5823-1985; Simonds, Erin/0000-0002-3497-4861; Pe'er, Dana/0000-0002-9259-8817	Damon Runyon Cancer Research Foundation [DRG-2017-09]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [PN2EY018228, 0158 G KB065, U19 AI057229, P01 CA034233-22A1, HHSN272200700038C, 1R01CA130826, RFA CA 09-011, RFA CA 09-009]; CIRMCalifornia Institute for Regenerative Medicine [USC DR1-01477]	Grant sponsor: Damon Runyon Cancer Research Foundation Fellowship; Grant number: DRG-2017-09; Grant sponsor: NIH; Grant numbers: PN2EY018228, 0158 G KB065, U19 AI057229, P01 CA034233-22A1, HHSN272200700038C, 1R01CA130826, RFA CA 09-011, and RFA CA 09-009; Grant sponsor: CIRM grant; Grant number: USC DR1-01477.	Abdelrahman AI, 2010, J ANAL ATOM SPECTROM, V25, P260, DOI 10.1039/b921770c; Abdelrahman AI, 2009, J AM CHEM SOC, V131, P15276, DOI 10.1021/ja9052009; Bandura DR, 2009, ANAL CHEM, V81, P6813, DOI 10.1021/ac901049w; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Bodenmiller B, 2012, NAT BIOTECHNOL, V30, P858, DOI 10.1038/nbt.2317; Dendrou Calliope A., 2009, Advances in Bioinformatics, V2009, P476106, DOI 10.1155/2009/476106; Hahne F, 2010, CYTOM PART A, V77A, P121, DOI 10.1002/cyto.a.20823; Ornatsky O, 2010, J IMMUNOL METHODS, V361, P1, DOI 10.1016/j.jim.2010.07.002; Perfetto SP, 2006, NAT PROTOC, V1, P1522, DOI 10.1038/nprot.2006.250	9	254	253	6	51	WILEY	HOBOKEN	111 RIVER ST, HOBOKEN 07030-5774, NJ USA	1552-4922	1552-4930		CYTOM PART A	Cytom. Part A	MAY	2013	83A	5					483	+		10.1002/cyto.a.22271			12	Biochemical Research Methods; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	133CW	WOS:000318116000001	23512433	Green Accepted			2020-07-01	
J	Horowitz, A; Strauss-Albee, DM; Leipold, M; Kubo, J; Nemat-Gorgani, N; Dogan, OC; Dekker, CL; Mackey, S; Maecker, H; Swan, GE; Davis, MM; Norman, PJ; Guethlein, LA; Desai, M; Parham, P; Blish, CA				Horowitz, Amir; Strauss-Albee, Dara M.; Leipold, Michael; Kubo, Jessica; Nemat-Gorgani, Neda; Dogan, Ozge C.; Dekker, Cornelia L.; Mackey, Sally; Maecker, Holden; Swan, Gary E.; Davis, Mark M.; Norman, Paul J.; Guethlein, Lisbeth A.; Desai, Manisha; Parham, Peter; Blish, Catherine A.			Genetic and Environmental Determinants of Human NK Cell Diversity Revealed by Mass Cytometry	SCIENCE TRANSLATIONAL MEDICINE			English	Article							NATURAL-KILLER-CELLS; HLA-C; INHIBITORY RECEPTORS; CYTOMEGALOVIRUS-INFECTION; MISSING SELF; SPECIFICITY; REPERTOIRES; ACTIVATION; KIR; CYTOTOXICITY	Natural killer (NK) cells play critical roles in immune defense and reproduction, yet remain the most poorly understood major lymphocyte population. Because their activation is controlled by a variety of combinatorially expressed activating and inhibitory receptors, NK cell diversity and function are closely linked. To provide an unprecedented understanding of NK cell repertoire diversity, we used mass cytometry to simultaneously analyze 37 parameters, including 28 NK cell receptors, on peripheral blood NK cells from 5 sets of monozygotic twins and 12 unrelated donors of defined human leukocyte antigen (HLA) and killer cell immunoglobulin-like receptor (KIR) genotype. This analysis revealed a remarkable degree of NK cell diversity, with an estimated 6000 to 30,000 phenotypic populations within an individual and >100,000 phenotypes in the donor panel. Genetics largely determined inhibitory receptor expression, whereas activation receptor expression was heavily environmentally influenced. Therefore, NK cells may maintain self-tolerance through strictly regulated expression of inhibitory receptors while using adaptable expression patterns of activating and costimulatory receptors to respond to pathogens and tumors. These findings further suggest the possibility that discrete NK cell subpopulations could be harnessed for immunotherapeutic strategies in the settings of infection, reproduction, and transplantation.	[Horowitz, Amir; Nemat-Gorgani, Neda; Norman, Paul J.; Guethlein, Lisbeth A.; Parham, Peter] Stanford Univ, Sch Med, Dept Biol Struct, Stanford, CA 94305 USA; [Horowitz, Amir; Leipold, Michael; Maecker, Holden; Davis, Mark M.; Parham, Peter] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Horowitz, Amir; Strauss-Albee, Dara M.; Davis, Mark M.; Parham, Peter; Blish, Catherine A.] Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA; [Strauss-Albee, Dara M.; Kubo, Jessica; Dogan, Ozge C.; Desai, Manisha; Blish, Catherine A.] Stanford Univ, Sch Med, Dept Med, Stanford, CA 94305 USA; [Dekker, Cornelia L.; Mackey, Sally] Stanford Univ, Sch Med, Dept Pediat, Stanford, CA 94305 USA; [Swan, Gary E.] SRI Int, Ctr Hlth Sci, Menlo Pk, CA 94025 USA[Horowitz, Amir; Nemat-Gorgani, Neda; Norman, Paul J.; Guethlein, Lisbeth A.; Parham, Peter] Stanford Univ, Sch Med, Dept Biol Struct, Stanford, CA 94305 USA; [Horowitz, Amir; Leipold, Michael; Maecker, Holden; Davis, Mark M.; Parham, Peter] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Horowitz, Amir; Strauss-Albee, Dara M.; Davis, Mark M.; Parham, Peter; Blish, Catherine A.] Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA; [Strauss-Albee, Dara M.; Kubo, Jessica; Dogan, Ozge C.; Desai, Manisha; Blish, Catherine A.] Stanford Univ, Sch Med, Dept Med, Stanford, CA 94305 USA; [Dekker, Cornelia L.; Mackey, Sally] Stanford Univ, Sch Med, Dept Pediat, Stanford, CA 94305 USA; [Swan, Gary E.] SRI Int, Ctr Hlth Sci, Menlo Pk, CA 94025 USA	Blish, CA (reprint author), Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA.	cblish@stanford.educblish@stanford.edu	Leipold, Michael/A-9452-2013	Leipold, Michael/0000-0001-5389-0906; Blish, Catherine/0000-0001-6946-7627; Strauss-Albee, Dara/0000-0003-1235-7800; Norman, Paul/0000-0001-8370-7703	NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [T32 AI07290, AI22039, U19AI090019, U19 AI057229]; NSFNational Science Foundation (NSF) [DGE-114740]; NIH/National Center for Research Resources Clinical and Translational Science Award [UL1 RR025744]; University of Washington Center for AIDS Research [P30 AI027757]; Beckman Young Investigator Award	This study was funded by NIH training grant T32 AI07290 (A. H.), NSF training grant DGE-114740 (D. M. S.-A.), NIH grants AI22039 (P. P.) and U19AI090019 and U19 AI057229 (M. M. D.), NIH/National Center for Research Resources Clinical and Translational Science Award UL1 RR025744, a New Investigator Award from the University of Washington Center for AIDS Research (P30 AI027757, C. A. B.), and a Beckman Young Investigator Award (C.A.B.).	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Transl. Med.	OCT 23	2013	5	208							208ra145	10.1126/scitranslmed.3006702			11	Cell Biology; Medicine, Research & Experimental	Cell Biology; Research & Experimental Medicine	240IX	WOS:000326090200006	24154599	Green Accepted			2020-07-01	
J	Chevrier, S; Levine, JH; Zanotelli, VRT; Silina, K; Schulz, D; Bacac, M; Ries, CH; Ailles, L; Jewett, MAS; Moch, H; van den Broek, M; Beisel, C; Stadler, MB; Gedye, C; Reis, B; Pe'er, D; Bodenmiller, B				Chevrier, Stephane; Levine, Jacob Harrison; Zanotelli, Vito Riccardo Tomaso; Silina, Karina; Schulz, Daniel; Bacac, Marina; Ries, Carola Hermine; Ailles, Laurie; Jewett, Michael Alexander Spencer; Moch, Holger; van den Broek, Maries; Beisel, Christian; Stadler, Michael Beda; Gedye, Craig; Reis, Bernhard; Pe'er, Dana; Bodenmiller, Bernd			An Immune Atlas of Clear Cell Renal Cell Carcinoma	CELL			English	Article							TUMOR-ASSOCIATED MACROPHAGES; T-CELLS; GLIOMA PROGRESSION; CANCER; POLARIZATION; EXPRESSION; ACTIVATION; REVEALS; CD38; MECHANISMS	Immune cells in the tumor microenvironment modulate cancer progression and are attractive therapeutic targets. Macrophages and T cells are key components of the microenvironment, yet their phenotypes and relationships in this ecosystem and to clinical outcomes are ill defined. We used mass cytometry with extensive antibody panels to perform in-depth immune profiling of samples from 73 clear cell renal cell carcinoma (ccRCC) patients and five healthy controls. In 3.5 million measured cells, we identified 17 tumor-associated macrophage phenotypes, 22 T cell phenotypes, and a distinct immune composition correlated with progression-free survival, thereby presenting an in-depth human atlas of the immune tumor microenvironment in this disease. This study revealed potential biomarkers and targets for immunotherapy development and validated tools that can be used for immune profiling of other tumor types.	[Chevrier, Stephane; Zanotelli, Vito Riccardo Tomaso; Schulz, Daniel; Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, Winterthurerstr 190, CH-8057 Zurich, Switzerland; [Levine, Jacob Harrison; Pe'er, Dana] Sloan Kettering Inst, Computat & Syst Biol Program, 1275 York Ave, New York, NY 10065 USA; [Zanotelli, Vito Riccardo Tomaso] ETH, Syst Biol PhD Program, Life Sci Zurich Grad Sch, CH-8057 Zurich, Switzerland; [Zanotelli, Vito Riccardo Tomaso] Univ Zurich, CH-8057 Zurich, Switzerland; [Silina, Karina; van den Broek, Maries] Univ Zurich, Inst Expt Immunol, Winterthurerstr 190, CH-8057 Zurich, Switzerland; [Bacac, Marina] Roche Innovat Ctr Zurich, Roche Pharma Res & Early Dev, Wagistr 18, CH-8952 Schlieren, Switzerland; [Ries, Carola Hermine] Roche Innovat Ctr Zurich, Roche Pharma Res & Early Dev, Nonnenwald 2, D-82377 Penzberg, Germany; [Ailles, Laurie] Univ Toronto, Dept Med Biophys, Toronto, ON M5G 1L7, Canada; [Ailles, Laurie; Jewett, Michael Alexander Spencer] Univ Hlth Network, Princess Margaret Canc Ctr, Toronto, ON M5G 1L7, Canada; [Moch, Holger] Univ Hosp Zurich, Inst Surg Pathol, Schmelzbergstr 12, CH-8091 Zurich, Switzerland; [Beisel, Christian] ETH, Dept Biosyst Sci & Engn, Mattenstr 26, CH-4058 Basel, Switzerland; [Stadler, Michael Beda] Friedrich Miescher Inst Biomed Res, Maulbeerstr 66, CH-4058 Basel, Switzerland; [Stadler, Michael Beda] Swiss Inst Bioinformat, Mattenstr 26, CH-4058 Basel, Switzerland; [Gedye, Craig] Univ Newcastle, Hunter Med Res Inst, Sch Biomed Sci & Pharm, Newcastle, NSW 2035, Australia; [Reis, Bernhard] Roche Innovat Ctr Basel, Pharmaceut Sci, Roche Pharma Res & Early Dev, Grenzacherstr 124, CH-4070 Basel, Switzerland[Chevrier, Stephane; Zanotelli, Vito Riccardo Tomaso; Schulz, Daniel; Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, Winterthurerstr 190, CH-8057 Zurich, Switzerland; [Levine, Jacob Harrison; Pe'er, Dana] Sloan Kettering Inst, Computat & Syst Biol Program, 1275 York Ave, New York, NY 10065 USA; [Zanotelli, Vito Riccardo Tomaso] ETH, Syst Biol PhD Program, Life Sci Zurich Grad Sch, CH-8057 Zurich, Switzerland; [Zanotelli, Vito Riccardo Tomaso] Univ Zurich, CH-8057 Zurich, Switzerland; [Silina, Karina; van den Broek, Maries] Univ Zurich, Inst Expt Immunol, Winterthurerstr 190, CH-8057 Zurich, Switzerland; [Bacac, Marina] Roche Innovat Ctr Zurich, Roche Pharma Res & Early Dev, Wagistr 18, CH-8952 Schlieren, Switzerland; [Ries, Carola Hermine] Roche Innovat Ctr Zurich, Roche Pharma Res & Early Dev, Nonnenwald 2, D-82377 Penzberg, Germany; [Ailles, Laurie] Univ Toronto, Dept Med Biophys, Toronto, ON M5G 1L7, Canada; [Ailles, Laurie; Jewett, Michael Alexander Spencer] Univ Hlth Network, Princess Margaret Canc Ctr, Toronto, ON M5G 1L7, Canada; [Moch, Holger] Univ Hosp Zurich, Inst Surg Pathol, Schmelzbergstr 12, CH-8091 Zurich, Switzerland; [Beisel, Christian] ETH, Dept Biosyst Sci & Engn, Mattenstr 26, CH-4058 Basel, Switzerland; [Stadler, Michael Beda] Friedrich Miescher Inst Biomed Res, Maulbeerstr 66, CH-4058 Basel, Switzerland; [Stadler, Michael Beda] Swiss Inst Bioinformat, Mattenstr 26, CH-4058 Basel, Switzerland; [Gedye, Craig] Univ Newcastle, Hunter Med Res Inst, Sch Biomed Sci & Pharm, Newcastle, NSW 2035, Australia; [Reis, Bernhard] Roche Innovat Ctr Basel, Pharmaceut Sci, Roche Pharma Res & Early Dev, Grenzacherstr 124, CH-4070 Basel, Switzerland	Bodenmiller, B (reprint author), Univ Zurich, Inst Mol Life Sci, Winterthurerstr 190, CH-8057 Zurich, Switzerland.	bernd.bodenmiller@imls.uzh.chbernd.bodenmiller@imls.uzh.ch	van den Broek, Maries/L-6532-2019; wang, edwin/G-5372-2010	Gedye, Craig/0000-0002-1629-4352; Zanotelli, Vito Riccardo Tomaso/0000-0001-7268-311X; van den Broek, Maries/0000-0002-9489-3692; Pe'er, Dana/0000-0002-9259-8817; Reis, Bernhard/0000-0001-7611-8222	SNSFSwiss National Science Foundation (SNSF); SystemsX Transfer Project "Friends and Foes"; SystemsX MetastasiX grant; European Research Council (ERC) under the European Union/ERCEuropean Research Council (ERC) [336921]; RocheRoche Holding; EMBO - European Commission [ALTF 970-2014, GA-2013-609409]; RACP CSL Fellowship; CIHR/KCC SHOPP Fellowship; NHMRCNational Health and Medical Research Council of Australia [628939]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DP1-HD084071, R01CA164729]; SNSFSwiss National Science Foundation (SNSF) [31003A_152851]; Zurich Cancer League grant; Stiftung fur Forschung in der Onkologie grant; University Research Priority Project "Translational Cancer Research" grant; MSK Cancer Center [P30 CA008748]; F. Hoffmann-La Roche AGHoffmann-La Roche	We are very grateful for the generous donation of tumor samples by patients undergoing surgery that was collected through the University Health Network Biobank and Cooperative Health Tissue Network. We thank Manfred Kopf, Luigi Tortola, Sabine Hoves, Sylvia Herter, and the Bodenmiller lab for fruitful discussions; Andrew Evans for pathological analysis of tissue samples; Andrea Jacobs and Stefanie Engler for technical support; Christina Ewald for cell sorting; the CyTOF facility and the Center for Microscopy and Image Analysis (University of Zurich); and the Genomics Facility Basel. B.B.'s research is funded by a SNSF R'Equip grant, a SNSF Assistant Professorship grant, the SystemsX Transfer Project "Friends and Foes," the SystemsX MetastasiX grant, and the European Research Council (ERC) under the European Union's Seventh Framework Program (FP/2007-2013/ERC grant agreement no. 336921). This work was funded by a Roche Postdoctoral Fellowship (to S.C.); an EMBO fellowship (ALTF 970-2014) co-funded by the European Commission (LTFCOFUND2013, GA-2013-609409) (to D.S.); an RACP CSL Fellowship (to C.G.); a CIHR/KCC SHOPP Fellowship (to C.G.), an NHMRC Early Career Fellowship (628939 to C.G.), NIH grants (DP1-HD084071 and R01CA164729), a SNSF grant (31003A_152851 to M.v.d.B.), a Zurich Cancer League grant (to M.v.d.B. and K.S.), a Stiftung fur Forschung in der Onkologie grant (to M.v.d.B.), a University Research Priority Project "Translational Cancer Research" grant (to M.v.d.B.), and a MSK Cancer Center Support Grant/Core Grant (P30 CA008748) (to D.P.). This work was in part supported by a fellowship from F. Hoffmann-La Roche AG. M.B., C.R., and B.R. are employees for F. Hoffmann-La Roche AG.	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J	Spitzer, MH; Carmi, Y; Reticker-Flynn, NE; Kwek, SS; Madhireddy, D; Martins, MM; Gherardini, PF; Prestwood, TR; Chabon, J; Bendall, SC; Fong, L; Nolan, GP; Engleman, EG				Spitzer, Matthew H.; Carmi, Yaron; Reticker-Flynn, Nathan E.; Kwek, Serena S.; Madhireddy, Deepthi; Martins, Maria M.; Gherardini, Pier Federico; Prestwood, Tyler R.; Chabon, Jonathan; Bendall, Sean C.; Fong, Lawrence; Nolan, Garry P.; Engleman, Edgar G.			Systemic Immunity Is Required for Effective Cancer Immunotherapy	CELL			English	Article							REGULATORY T-CELLS; MASS CYTOMETRY; BREAST-CANCER; TUMOR; IDENTIFICATION; PATIENT	Immune responses involve coordination across cell types and tissues. However, studies in cancer immunotherapy have focused heavily on local immune responses in the tumor microenvironment. To investigate immune activity more broadly, we performed an organism-wide study in genetically engineered cancer models using mass cytometry. We analyzed immune responses in several tissues after immunotherapy by developing intuitive models for visualizing single-cell data with statistical inference. Immune activation was evident in the tumor and systemically shortly after effective therapy was administered. However, during tumor rejection, only peripheral immune cells sustained their proliferation. This systemic response was coordinated across tissues and required for tumor eradication in several immunotherapy models. An emergent population of peripheral CD4 T cells conferred protection against new tumors and was significantly expanded in patients responding to immunotherapy. These studies demonstrate the critical impact of systemic immune responses that drive tumor rejection.	[Spitzer, Matthew H.; Carmi, Yaron; Reticker-Flynn, Nathan E.; Martins, Maria M.; Prestwood, Tyler R.; Chabon, Jonathan; Bendall, Sean C.; Engleman, Edgar G.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Madhireddy, Deepthi; Gherardini, Pier Federico; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Nolan, Garry P.; Engleman, Edgar G.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Kwek, Serena S.; Fong, Lawrence] Univ Calif San Francisco, Dept Med, Div Hematol & Oncol, San Francisco, CA 94143 USA; [Spitzer, Matthew H.; Fong, Lawrence] Univ Calif San Francisco, Helen Diller Comprehens Canc Ctr, San Francisco, CA 94143 USA; [Carmi, Yaron] Tel Aviv Univ, Sackler Sch Med, Dept Pathol, IL-69978 Ramat Aviv, Israel[Spitzer, Matthew H.; Carmi, Yaron; Reticker-Flynn, Nathan E.; Martins, Maria M.; Prestwood, Tyler R.; Chabon, Jonathan; Bendall, Sean C.; Engleman, Edgar G.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Madhireddy, Deepthi; Gherardini, Pier Federico; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Nolan, Garry P.; Engleman, Edgar G.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Kwek, Serena S.; Fong, Lawrence] Univ Calif San Francisco, Dept Med, Div Hematol & Oncol, San Francisco, CA 94143 USA; [Spitzer, Matthew H.; Fong, Lawrence] Univ Calif San Francisco, Helen Diller Comprehens Canc Ctr, San Francisco, CA 94143 USA; [Carmi, Yaron] Tel Aviv Univ, Sackler Sch Med, Dept Pathol, IL-69978 Ramat Aviv, Israel	Spitzer, MH; Engleman, EG (reprint author), Stanford Univ, Dept Pathol, Stanford, CA 94305 USA.; Spitzer, MH; Nolan, GP (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.; Spitzer, MH; Nolan, GP; Engleman, EG (reprint author), Stanford Univ, Program Immunol, Stanford, CA 94305 USA.; Spitzer, MH (reprint author), Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA.; Spitzer, MH (reprint author), Univ Calif San Francisco, Helen Diller Comprehens Canc Ctr, San Francisco, CA 94143 USA.	matthew.spitzer@ucsf.edu; gnolan@stanford.edu; edengleman@stanford.edumatthew.spitzer@ucsf.edu; gnolan@stanford.edu; edengleman@stanford.edu	Reticker-Flynn, Nathan/R-7226-2019; Nolan, Garry/AAE-7903-2019	Reticker-Flynn, Nathan/0000-0002-9963-039X; Spitzer, Matthew/0000-0002-5291-3819	NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [F31CA189331, DP5OD023056, F32CA189408, U19AI057229, U19AI100627, R33CA183654, R01HL120724, R01CA196657, U54CA209971, R01AI118884]; DODUnited States Department of Defense [OC110674, 11491122]; Gates FoundationGates Foundation [OPP1113682]; NIAIDUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [HHSN272201200028C]; FDAUnited States Department of Health & Human Services [HHSF223201210194C BAA-12-00118]	We thank G.K. Fragiadakis and L.E. Sanman for helpful conversations and feedback and A. Trejo and A. Jager for CyTOF maintenance. M.H.S., P.F.G., L.F., and G.P.N. are investigators of the Parker Institute for Cancer Immunotherapy. G.P.N. has a personal financial interest in, and S.C.B. has been a paid consultant for, Fluidigm, manufacturer of the mass cytometer utilized. E.G.E. is a founder and board member of Bolt Biotherapeutics, licensee of the therapy with alloIgG. This work was supported by NIH grants F31CA189331 and DP5OD023056 to M.H.S.; NIH grant F32CA189408 to N.R.F.; NIH grants U19AI057229, U19AI100627, R33CA183654, and R01HL120724, DOD grants OC110674 and 11491122, Gates Foundation grant OPP1113682, NIAID grant HHSN272201200028C, and FDA grant HHSF223201210194C BAA-12-00118 to G.P.N.; NIH grants R01CA196657 and U54CA209971 to G.P.N. and E.G.E.; and NIH grant R01AI118884 to E.G.E.	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J	Setty, M; Tadmor, MD; Reich-Zeliger, S; Ange, O; Salame, TM; Kathail, P; Choi, K; Bendall, S; Friedman, N; Pe'er, D				Setty, Manu; Tadmor, Michelle D.; Reich-Zeliger, Shlomit; Ange, Omer; Salame, Tomer Meir; Kathail, Pooja; Choi, Kristy; Bendall, Sean; Friedman, Nir; Pe'er, Dana			Wishbone identifies bifurcating developmental trajectories from single-cell data	NATURE BIOTECHNOLOGY			English	Article							GENE-EXPRESSION; TRANSCRIPTION FACTORS; DIFFUSION MAPS; RNA-SEQ; DIFFERENTIATION; TRANSFORMATION; HETEROGENEITY; MECHANISMS; LANDSCAPE; NETWORKS	Recent single-cell analysis technologies offer an unprecedented opportunity to elucidate developmental pathways. Here we present Wishbone, an algorithm for positioning single cells along bifurcating developmental trajectories with high resolution. Wishbone uses multi-dimensional single-cell data, such as mass cytometry or RNA-Seq data, as input and orders cells according to their developmental progression, and it pinpoints bifurcation points by labeling each cell as pre-bifurcation or as one of two post-bifurcation cell fates. Using 30-channel mass cytometry data, we show that Wishbone accurately recovers the known stages of T-cell development in the mouse thymus, including the bifurcation point. We also apply the algorithm to mouse myeloid differentiation and demonstrate its generalization to additional lineages. A comparison of Wishbone to diffusion maps, SCUBA and Monocle shows that it outperforms these methods both in the accuracy of ordering cells and in the correct identification of branch points.	[Setty, Manu; Tadmor, Michelle D.; Kathail, Pooja; Choi, Kristy; Pe'er, Dana] Columbia Univ, Dept Syst Biol, Dept Biol Sci, New York, NY USA; [Reich-Zeliger, Shlomit; Friedman, Nir] Weizmann Inst Sci, Dept Immunol, IL-76100 Rehovot, Israel; [Ange, Omer] Univ British Columbia, Dept Math, Vancouver, BC, Canada; [Salame, Tomer Meir] Weizmann Inst Sci, Biol Serv Unit, IL-76100 Rehovot, Israel; [Bendall, Sean] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA[Setty, Manu; Tadmor, Michelle D.; Kathail, Pooja; Choi, Kristy; Pe'er, Dana] Columbia Univ, Dept Syst Biol, Dept Biol Sci, New York, NY USA; [Reich-Zeliger, Shlomit; Friedman, Nir] Weizmann Inst Sci, Dept Immunol, IL-76100 Rehovot, Israel; [Ange, Omer] Univ British Columbia, Dept Math, Vancouver, BC, Canada; [Salame, Tomer Meir] Weizmann Inst Sci, Biol Serv Unit, IL-76100 Rehovot, Israel; [Bendall, Sean] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA	Pe'er, D (reprint author), Columbia Univ, Dept Syst Biol, Dept Biol Sci, New York, NY USA.	dpeer@biology.columbia.edudpeer@biology.columbia.edu		Pe'er, Dana/0000-0002-9259-8817; Setty, Manu/0000-0002-0344-2627	NSFNational Science Foundation (NSF); NSFNational Science Foundation (NSF) [MCB-1149728]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DP1-HD084071, R01CA164729]; David and Fela Shapell Family Foundation INCPM Fund; Nissim Center, for the Development of Scientific Resources; ISFIsrael Science Foundation [1184/15]	We would like to thank A. Bloemendal, Z. Good, N. Hacohen, S. Krishnaswamy, J. Levine and A.J. Carr for their helpful comments. M.D.T. is supported by an NSF graduate fellowship. This work was supported by NSF MCB-1149728, NIH DP1-HD084071, NIH R01CA164729 to D.P. D.P. holds a Packard Fellowship for Science and Engineering. This work was also supported by David and Fela Shapell Family Foundation INCPM Fund, the WIS staff scientists grant from the Nissim Center, for the Development of Scientific Resources, and ISF 1184/15 to N.F.	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Biotechnol.	JUN	2016	34	6					637	645		10.1038/nbt.3569			9	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	DO5TT	WOS:000377846400031	27136076	Green Accepted			2020-07-01	
J	Rao, DA; Gurish, MF; Marshall, JL; Slowikowski, K; Fonseka, CY; Liu, YY; Donlin, LT; Henderson, LA; Wei, K; Mizoguchi, F; Teslovich, NC; Weinblatt, ME; Massarotti, EM; Coblyn, JS; Helfgott, SM; Lee, YC; Todd, DJ; Ykerk, VPB; Goodman, SM; Pernis, AB; Ivashkiv, LB; Karlson, EW; Nigrovic, PA; Filer, A; Buckley, CD; Lederer, JA; Raychaudhuri, S; Renner, MBB				Rao, Deepak A.; Gurish, Michael F.; Marshall, Jennifer L.; Slowikowski, Kamil; Fonseka, Chamith Y.; Liu, Yanyan; Donlin, Laura T.; Henderson, Lauren A.; Wei, Kevin; Mizoguchi, Fumitaka; Teslovich, Nikola C.; Weinblatt, Michael E.; Massarotti, Elena M.; Coblyn, Jonathan S.; Helfgott, Simon M.; Lee, Yvonne C.; Todd, Derrick J.; Ykerk, Vivian P. B.; Goodman, Susan M.; Pernis, Alessandra B.; Ivashkiv, Lionel B.; Karlson, Elizabeth W.; Nigrovic, Peter A.; Filer, Andrew; Buckley, Christopher D.; Lederer, James A.; Raychaudhuri, Soumya; Renner, Michael B. B.			Pathologically expanded peripheral T helper cell subset drives B cells in rheumatoid arthritis	NATURE			English	Article							FOLLICULAR HELPER; EXTRAVASATION; CXCL13; BCL6	CD4(+) T cells are central mediators of autoimmune pathology; however, defining their key effector functions in specific autoimmune diseases remains challenging. Pathogenic CD4(+) T cells within affected tissues may be identified by expression of markers of recent activation(1). Here we use mass cytometry to analyse activated T cells in joint tissue from patients with rheumatoid arthritis, a chronic immune-mediated arthritis that affects up to 1% of the population(2). This approach revealed a markedly expanded population of PD-1(hi)CXCR5(-)CD4(+) T cells in synovium of patients with rheumatoid arthritis. However, these cells are not exhausted, despite high PD-1 expression. Rather, using multidimensional cytometry, transcriptomics, and functional assays, we define a population of PD-1(hi)CXCR5(-) 'peripheral helper' T (T-PH) cells that express factors enabling B-cell help, including IL-21, CXCL13, ICOS, and MAF. Like PD-1(hi)CXCR5(+) T follicular helper cells, T-PH cells induce plasma cell differentiation in vitro through IL-21 secretion and SLAMF5 interaction (refs 3, 4). However, global transcriptomics highlight differences between T-PH cells and T follicular helper cells, including altered expression of BCL6 and BLIMP1 and unique expression of chemokine receptors that direct migration to inflamed sites, such as CCR2, CX3CR1, and CCR5, in T-PH cells. T-PH cells appear to be uniquely poised to promote B-cell responses and antibody production within pathologically inflamed non-lymphoid tissues.	[Rao, Deepak A.; Gurish, Michael F.; Slowikowski, Kamil; Fonseka, Chamith Y.; Liu, Yanyan; Henderson, Lauren A.; Wei, Kevin; Mizoguchi, Fumitaka; Teslovich, Nikola C.; Weinblatt, Michael E.; Karlson, Elizabeth W.] Brigham & Womens Hosp, Div Rheumatol Immunol & Allergy, Boston, MA 02115 USA; [Wei, Kevin; Todd, Derrick J.; Goodman, Susan M.] Harvard Med Sch, Boston, MA 02115 USA; [Marshall, Jennifer L.; Henderson, Lauren A.; Helfgott, Simon M.; Todd, Derrick J.] Univ Birmingham, Queen Elizabeth Hosp, Inst Inflammat & Ageing, Rheumatol Res Grp, Birmingham B15 2WB, W Midlands, England; [Gurish, Michael F.; Liu, Yanyan; Lee, Yvonne C.; Goodman, Susan M.] Brigham & Womens Hosp, Div Genet, Boston, MA 02115 USA; [Marshall, Jennifer L.; Donlin, Laura T.; Teslovich, Nikola C.; Helfgott, Simon M.; Lee, Yvonne C.] Harvard Med Sch, Boston, MA 02115 USA; [Slowikowski, Kamil; Todd, Derrick J.; Karlson, Elizabeth W.] MIT, Broad Inst, Program Med & Populat Genet, Cambridge, MA 02138 USA; [Rao, Deepak A.; Helfgott, Simon M.; Ykerk, Vivian P. B.; Karlson, Elizabeth W.] Harvard Univ, Cambridge, MA 02138 USA; [Fonseka, Chamith Y.] Partners Ctr Personalized Genet Med, Boston, MA 02115 USA; [Donlin, Laura T.; Henderson, Lauren A.] Harvard Univ, Bioinformat & Integrat Genom, Cambridge, MA 02138 USA; [Helfgott, Simon M.] Harvard Univ, Biol & Biomed Sci, Cambridge, MA 02138 USA; [Todd, Derrick J.; Karlson, Elizabeth W.] Hosp Special Surg, Arthrit & Tissue Degenerat Program, New York, NY USA; [Helfgott, Simon M.; Ivashkiv, Lionel B.; Buckley, Christopher D.; Raychaudhuri, Soumya] Hosp Special Surg, David Z Rosensweig Genom Res Ctr, New York, NY 10021 USA; [Marshall, Jennifer L.; Wei, Kevin; Coblyn, Jonathan S.; Lee, Yvonne C.; Ivashkiv, Lionel B.] Boston Childrens Hosp, Divis Immunol, Boston, MA 02115 USA; [Coblyn, Jonathan S.; Pernis, Alessandra B.; Nigrovic, Peter A.] Hosp Special Surg, Div Rheumatol, 535 E 70th St, New York, NY 10021 USA; [Slowikowski, Kamil; Mizoguchi, Fumitaka; Massarotti, Elena M.; Goodman, Susan M.] Cornell Univ, Weill Cornell Med Coll, New York, NY 10021 USA; [Massarotti, Elena M.; Lee, Yvonne C.; Ykerk, Vivian P. B.; Pernis, Alessandra B.] Hosp Special Surg, Autoimmun & Inflammat Program, New York, NY 10021 USA; [Donlin, Laura T.; Goodman, Susan M.] Brigham & Womens Hosp, Dept Surg, Boston, MA 02115 USA; [Henderson, Lauren A.; Weinblatt, Michael E.; Goodman, Susan M.] Karolinska Inst, Reumatol Unit, S-17176 Stockholm, Sweden; [Mizoguchi, Fumitaka; Massarotti, Elena M.; Lee, Yvonne C.; Todd, Derrick J.] Karolinska Univ Hosp Solna, S-17176 Stockholm, Sweden; [Massarotti, Elena M.; Todd, Derrick J.; Ivashkiv, Lionel B.] Univ Manchester, Inst Inflammat & Repair, Manchester M13 9PT, Lancs, England[Rao, Deepak A.; Gurish, Michael F.; Slowikowski, Kamil; Fonseka, Chamith Y.; Liu, Yanyan; Henderson, Lauren A.; Wei, Kevin; Mizoguchi, Fumitaka; Teslovich, Nikola C.; Weinblatt, Michael E.; Karlson, Elizabeth W.] Brigham & Womens Hosp, Div Rheumatol Immunol & Allergy, Boston, MA 02115 USA; [Wei, Kevin; Todd, Derrick J.; Goodman, Susan M.] Harvard Med Sch, Boston, MA 02115 USA; [Marshall, Jennifer L.; Henderson, Lauren A.; Helfgott, Simon M.; Todd, Derrick J.] Univ Birmingham, Queen Elizabeth Hosp, Inst Inflammat & Ageing, Rheumatol Res Grp, Birmingham B15 2WB, W Midlands, England; [Gurish, Michael F.; Liu, Yanyan; Lee, Yvonne C.; Goodman, Susan M.] Brigham & Womens Hosp, Div Genet, Boston, MA 02115 USA; [Marshall, Jennifer L.; Donlin, Laura T.; Teslovich, Nikola C.; Helfgott, Simon M.; Lee, Yvonne C.] Harvard Med Sch, Boston, MA 02115 USA; [Slowikowski, Kamil; Todd, Derrick J.; Karlson, Elizabeth W.] MIT, Broad Inst, Program Med & Populat Genet, Cambridge, MA 02138 USA; [Rao, Deepak A.; Helfgott, Simon M.; Ykerk, Vivian P. B.; Karlson, Elizabeth W.] Harvard Univ, Cambridge, MA 02138 USA; [Fonseka, Chamith Y.] Partners Ctr Personalized Genet Med, Boston, MA 02115 USA; [Donlin, Laura T.; Henderson, Lauren A.] Harvard Univ, Bioinformat & Integrat Genom, Cambridge, MA 02138 USA; [Helfgott, Simon M.] Harvard Univ, Biol & Biomed Sci, Cambridge, MA 02138 USA; [Todd, Derrick J.; Karlson, Elizabeth W.] Hosp Special Surg, Arthrit & Tissue Degenerat Program, New York, NY USA; [Helfgott, Simon M.; Ivashkiv, Lionel B.; Buckley, Christopher D.; Raychaudhuri, Soumya] Hosp Special Surg, David Z Rosensweig Genom Res Ctr, New York, NY 10021 USA; [Marshall, Jennifer L.; Wei, Kevin; Coblyn, Jonathan S.; Lee, Yvonne C.; Ivashkiv, Lionel B.] Boston Childrens Hosp, Divis Immunol, Boston, MA 02115 USA; [Coblyn, Jonathan S.; Pernis, Alessandra B.; Nigrovic, Peter A.] Hosp Special Surg, Div Rheumatol, 535 E 70th St, New York, NY 10021 USA; [Slowikowski, Kamil; Mizoguchi, Fumitaka; Massarotti, Elena M.; Goodman, Susan M.] Cornell Univ, Weill Cornell Med Coll, New York, NY 10021 USA; [Massarotti, Elena M.; Lee, Yvonne C.; Ykerk, Vivian P. B.; Pernis, Alessandra B.] Hosp Special Surg, Autoimmun & Inflammat Program, New York, NY 10021 USA; [Donlin, Laura T.; Goodman, Susan M.] Brigham & Womens Hosp, Dept Surg, Boston, MA 02115 USA; [Henderson, Lauren A.; Weinblatt, Michael E.; Goodman, Susan M.] Karolinska Inst, Reumatol Unit, S-17176 Stockholm, Sweden; [Mizoguchi, Fumitaka; Massarotti, Elena M.; Lee, Yvonne C.; Todd, Derrick J.] Karolinska Univ Hosp Solna, S-17176 Stockholm, Sweden; [Massarotti, Elena M.; Todd, Derrick J.; Ivashkiv, Lionel B.] Univ Manchester, Inst Inflammat & Repair, Manchester M13 9PT, Lancs, England	Rao, DA (reprint author), Brigham & Womens Hosp, Div Rheumatol Immunol & Allergy, Boston, MA 02115 USA.	darao@partners.org; mbrenner@research.bwh.harvard.edudarao@partners.org; mbrenner@research.bwh.harvard.edu	Mizoguchi, Fumitaka/Y-3271-2018; Donlin, Laura T/P-2483-2019; Slowikowski, Kamil/K-3728-2016	Mizoguchi, Fumitaka/0000-0002-3956-7122; Donlin, Laura T/0000-0002-1428-090X; Slowikowski, Kamil/0000-0002-2843-6370; Marshall, Jennifer/0000-0002-1727-8290; Raychaudhuri, Soumya/0000-0002-1901-8265; Rao, Deepak/0000-0001-9672-7746	William Docken Inflammatory Autoimmune Disease Fund; Mallinckrodt Research Fellowship; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [5U01GM092691-05, 1U19 AI111224-01]; Doris Duke Charitable FoundationDoris Duke Charitable Foundation (DDCF) [2013097]; Rheumatology Research Foundation Scientist Development Award; Arthritis Research UKVersus Arthritis [19791, 18547]; Fundacion Bechara;  [T32 AR007530-31];  [R01 AR064850-03];  [K01 AR066063];  [FP7-HEALTH-F2-2012-305549 EuroTEAM];  [P30 AR070253]; Arthritis Research UKVersus Arthritis [20088]	This work was supported by T32 AR007530-31 and the William Docken Inflammatory Autoimmune Disease Fund (to M.B.B.), Mallinckrodt Research Fellowship (to D.A.R.), R01 AR064850-03 (to Y.C.L.), NIH 5U01GM092691-05, 1U19 AI111224-01 and Doris Duke Charitable Foundation Grant #2013097 (to S.R.), Rheumatology Research Foundation Scientist Development Award (to L.A.H.), K01 AR066063 (to L.T.D.), Arthritis Research UK programme grant # 19791 (to C.D.B.), and Arthritis Research UK Clinician Scientist Fellowship # 18547 (to A.F.). J.L.M was supported by the FP7-HEALTH-F2-2012-305549 EuroTEAM. P.A.N. was supported by P30 AR070253 and the Fundacion Bechara. We thank A. Chicoine and the BWH Human Immunology Center Flow Cytometry Core for assistance with cell sorting.	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J	Becht, E; McInnes, L; Healy, J; Dutertre, CA; Kwok, IWH; Ng, LG; Ginhoux, F; Newell, EW				Becht, Etienne; McInnes, Leland; Healy, John; Dutertre, Charles-Antoine; Kwok, Immanuel W. H.; Ng, Lai Guan; Ginhoux, Florent; Newell, Evan W.			Dimensionality reduction for visualizing single-cell data using UMAP	NATURE BIOTECHNOLOGY			English	Article							ATLAS; CYTOMETRY	Advances in single-cell technologies have enabled high-resolution dissection of tissue composition. Several tools for dimensionality reduction are available to analyze the large number of parameters generated in single-cell studies. Recently, a nonlinear dimensionality-reduction technique, uniform manifold approximation and projection (UMAP), was developed for the analysis of any type of high-dimensional data. Here we apply it to biological data, using three well-characterized mass cytometry and single-cell RNA sequencing datasets. Comparing the performance of UMAP with five other tools, we find that UMAP provides the fastest run times, highest reproducibility and the most meaningful organization of cell clusters. The work highlights the use of UMAP for improved visualization and interpretation of single-cell data.	[Becht, Etienne; Dutertre, Charles-Antoine; Kwok, Immanuel W. H.; Ng, Lai Guan; Ginhoux, Florent; Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [McInnes, Leland; Healy, John] Tutte Inst Math & Comp, Ottawa, ON, Canada; [Newell, Evan W.] Fred Hutchinson Canc Res Ctr, Vaccine & Infect Dis Div, 1124 Columbia St, Seattle, WA 98104 USA[Becht, Etienne; Dutertre, Charles-Antoine; Kwok, Immanuel W. H.; Ng, Lai Guan; Ginhoux, Florent; Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [McInnes, Leland; Healy, John] Tutte Inst Math & Comp, Ottawa, ON, Canada; [Newell, Evan W.] Fred Hutchinson Canc Res Ctr, Vaccine & Infect Dis Div, 1124 Columbia St, Seattle, WA 98104 USA	Newell, EW (reprint author), ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore.; Newell, EW (reprint author), Fred Hutchinson Canc Res Ctr, Vaccine & Infect Dis Div, 1124 Columbia St, Seattle, WA 98104 USA.	enewell@fredhutch.orgenewell@fredhutch.org	Newell, Evan/AAE-9470-2020; Ng, Lai Guan/E-5673-2011	Newell, Evan/0000-0002-2889-243X; Ng, Lai Guan/0000-0003-1905-3586; Ginhoux, Florent/0000-0002-2857-7755; Dutertre, Charles-Antoine/0000-0001-7950-3652	A-STAR/SIgN core funding; A-STAR/SIgN immunomonitoring platform funding	We thank members of the Singapore Immunology Network and notably members of the E.W.N. laboratory. We thank S. Li, Y. Simoni, M. Chng, Y. Cheng, J. W. Lim and M. Fehlings for their insightful feedback. This study was funded by A-STAR/SIgN core funding and A-STAR/SIgN immunomonitoring platform funding.	Aibar S, 2017, NAT METHODS, V14, P1083, DOI [10.1038/NMETH.4463, 10.1038/nmeth.4463]; Amir ED, 2013, NAT BIOTECHNOL, V31, P545, DOI 10.1038/nbt.2594; Butler A, 2018, NAT BIOTECHNOL, V36, P411, DOI 10.1038/nbt.4096; Coifman RR, 2005, P NATL ACAD SCI USA, V102, P7426, DOI 10.1073/pnas.0500334102; De Graaf CA, 2016, STEM CELL REP, V7, P571, DOI 10.1016/j.stemcr.2016.07.007; Ding JR, 2018, NAT COMMUN, V9, DOI 10.1038/s41467-018-04368-5; Han XP, 2018, CELL, V172, P1091, DOI 10.1016/j.cell.2018.02.001; Huang H, 2016, SEMIN IMMUNOPATHOL, V38, P539, DOI 10.1007/s00281-016-0562-4; Levine JH, 2015, CELL, V162, P184, DOI 10.1016/j.cell.2015.05.047; Linderman G. C., 2017, PREPRINT; Martensson IL, 2007, CURR OPIN IMMUNOL, V19, P137, DOI 10.1016/j.coi.2007.02.006; McInnes L., 2018, PREPRINT; McInnes L, 2018, J OPEN SOURCE SOFTWA, V3, P861, DOI [10.21105/joss.00861, DOI 10.21105/joss.00861]; Saeys Y, 2016, NAT REV IMMUNOL, V16, P449, DOI 10.1038/nri.2016.56; Samusik N, 2016, NAT METHODS, V13, P493, DOI [10.1038/NMETH.3863, 10.1038/nmeth.3863]; Tenenbaum JB, 2000, SCIENCE, V290, P2319, DOI 10.1126/science.290.5500.2319; van der Maaten L, 2014, J MACH LEARN RES, V15, P3221; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; van Unen V, 2016, IMMUNITY, V44, P1227, DOI 10.1016/j.immuni.2016.04.014; Wattenberg M, 2016, DISTILL, DOI DOI 10.23915/DISTILL.00002; Wolf FA, 2018, GENOME BIOL, V19, DOI 10.1186/s13059-017-1382-0; Wong MT, 2016, IMMUNITY, V45, P442, DOI 10.1016/j.immuni.2016.07.007	22	208	208	35	75	NATURE PUBLISHING GROUP	NEW YORK	75 VARICK ST, 9TH FLR, NEW YORK, NY 10013-1917 USA	1087-0156	1546-1696		NAT BIOTECHNOL	Nat. Biotechnol.	JAN	2019	37	1					38	+		10.1038/nbt.4314			8	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	HG2PE	WOS:000454804600017	30531897				2020-07-01	
J	Chattopadhyay, PK; Gierahn, TM; Roederer, M; Love, JC				Chattopadhyay, Pratip K.; Gierahn, Todd M.; Roederer, Mario; Love, J. Christopher			Single-cell technologies for monitoring immune systems	NATURE IMMUNOLOGY			English	Review							NATURAL-KILLER-CELLS; FLOW-CYTOMETRY; MIGRATION BEHAVIOR; GENE-EXPRESSION; MASS CYTOMETRY; ACTIVATION; ANTIBODIES; SECRETION; HIERARCHY; RESPONSES	The complex heterogeneity of cells, and their interconnectedness with each other, are major challenges to identifying clinically relevant measurements that reflect the state and capability of the immune system. Highly multiplexed, single-cell technologies may be critical for identifying correlates of disease or immunological interventions as well as for elucidating the underlying mechanisms of immunity. Here we review limitations of bulk measurements and explore advances in single-cell technologies that overcome these problems by expanding the depth and breadth of functional and phenotypic analysis in space and time. The geometric increases in complexity of data make formidable hurdles for exploring, analyzing and presenting results. We summarize recent approaches to making such computations tractable and discuss challenges for integrating heterogeneous data obtained using these single-cell technologies.	[Chattopadhyay, Pratip K.; Roederer, Mario] NIAID, ImmunoTechnol Sect, Vaccine Res Ctr, NIH, Bethesda, MD 20892 USA; [Gierahn, Todd M.; Love, J. Christopher] MIT, Koch Inst Integrat Canc Res, Cambridge, MA 02139 USA[Chattopadhyay, Pratip K.; Roederer, Mario] NIAID, ImmunoTechnol Sect, Vaccine Res Ctr, NIH, Bethesda, MD 20892 USA; [Gierahn, Todd M.; Love, J. Christopher] MIT, Koch Inst Integrat Canc Res, Cambridge, MA 02139 USA	Love, JC (reprint author), MIT, Koch Inst Integrat Canc Res, 77 Massachusetts Ave, Cambridge, MA 02139 USA.	clove@mit.educlove@mit.edu		Chattopadhyay, Pratip/0000-0002-5457-9666	W.M. Keck FoundationW.M. Keck Foundation; US National Institute of Allergy And Infectious DiseasesUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [1U19AI089992, 1R56AI104274, 5R21AI106025]	This work was supported by the W.M. Keck Foundation and the US National Institute of Allergy And Infectious Diseases (1U19AI089992, 1R56AI104274 and 5R21AI106025). The content is solely the responsibility of the authors and does not necessarily represent the official views of the US National Institute of Allergy And Infectious Diseases or the US National Institutes of Health. We thank A. Shalek for helpful comments on scRNA-seq and N. Aghaeepour for discussions about data-analysis tools. J.C.L. is a Camille Dreyfus Teacher-Scholar. We acknowledge the service to the MIT community of the late Sean Collier.	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Immunol.	FEB	2014	15	2					128	135		10.1038/ni.2796			8	Immunology	Immunology	295XV	WOS:000330150600004	24448570	Green Accepted			2020-07-01	
J	Romee, R; Rosario, M; Berrien-Elliott, MM; Wagner, JA; Jewell, BA; Schappe, T; Leong, JW; Abdel-Latif, S; Schneider, SE; Willey, S; Neal, CC; Yu, LY; Oh, ST; Lee, YS; Mulder, A; Claas, F; Cooper, MA; Fehniger, TA				Romee, Rizwan; Rosario, Maximillian; Berrien-Elliott, Melissa M.; Wagner, Julia A.; Jewell, Brea A.; Schappe, Timothy; Leong, Jeffrey W.; Abdel-Latif, Sara; Schneider, Stephanie E.; Willey, Sarah; Neal, Carly C.; Yu, Liyang; Oh, Stephen T.; Lee, Yi-Shan; Mulder, Arend; Claas, Frans; Cooper, Megan A.; Fehniger, Todd A.			Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia	SCIENCE TRANSLATIONAL MEDICINE			English	Article							MASS CYTOMETRY; NK CELLS; TRANSPLANTATION; RECEPTOR; SURVIVAL; OUTCOMES; RELAPSE; IMMUNE; TREAT	Natural killer (NK) cells are an emerging cellular immunotherapy for patients with acute myeloid leukemia (AML); however, the best approach to maximize NK cell antileukemia potential is unclear. Cytokine-induced memory-like NK cells differentiate after a brief preactivation with interleukin-12 (IL-12), IL-15, and IL-18 and exhibit enhanced responses to cytokine or activating receptor restimulation for weeks to months after preactivation. We hypothesized that memory-like NK cells exhibit enhanced antileukemia functionality. We demonstrated that human memory-like NK cells have enhanced interferon-gamma production and cytotoxicity against leukemia cell lines or primary human AML blasts in vitro. Using mass cytometry, we found that memory-like NK cell functional responses were triggered against primary AML blasts, regardless of killer cell immunoglobulin-like receptor (KIR) to KIR-ligand interactions. In addition, multidimensional analyses identified distinct phenotypes of control and memory-like NK cells from the same individuals. Human memory-like NK cells xenografted into mice substantially reduced AML burden in vivo and improved overall survival. In the context of a first-in-human phase 1 clinical trial, adoptively transferred memory-like NK cells proliferated and expanded in AML patients and demonstrated robust responses against leukemia targets. Clinical responses were observed in five of nine evaluable patients, including four complete remissions. Thus, harnessing cytokine-induced memory-like NK cell responses represents a promising translational immunotherapy approach for patients with AML.	[Romee, Rizwan; Rosario, Maximillian; Berrien-Elliott, Melissa M.; Wagner, Julia A.; Jewell, Brea A.; Schappe, Timothy; Leong, Jeffrey W.; Abdel-Latif, Sara; Schneider, Stephanie E.; Willey, Sarah; Neal, Carly C.; Fehniger, Todd A.] Washington Univ, Sch Med, Dept Med, Div Oncol, St Louis, MO 63110 USA; [Rosario, Maximillian; Lee, Yi-Shan] Washington Univ, Sch Med, Dept Pathol, St Louis, MO 63110 USA; [Yu, Liyang; Oh, Stephen T.] Washington Univ, Sch Med, Dept Med, Div Hematol, St Louis, MO 63110 USA; [Mulder, Arend; Claas, Frans] Leiden Univ, Med Ctr, Dept Immunohematol & Blood Transfus, NL-2333 ZC Leiden, Netherlands; [Cooper, Megan A.] Washington Univ, Sch Med, Dept Pediat, Div Rheumatol, St Louis, MO 63110 USA[Romee, Rizwan; Rosario, Maximillian; Berrien-Elliott, Melissa M.; Wagner, Julia A.; Jewell, Brea A.; Schappe, Timothy; Leong, Jeffrey W.; Abdel-Latif, Sara; Schneider, Stephanie E.; Willey, Sarah; Neal, Carly C.; Fehniger, Todd A.] Washington Univ, Sch Med, Dept Med, Div Oncol, St Louis, MO 63110 USA; [Rosario, Maximillian; Lee, Yi-Shan] Washington Univ, Sch Med, Dept Pathol, St Louis, MO 63110 USA; [Yu, Liyang; Oh, Stephen T.] Washington Univ, Sch Med, Dept Med, Div Hematol, St Louis, MO 63110 USA; [Mulder, Arend; Claas, Frans] Leiden Univ, Med Ctr, Dept Immunohematol & Blood Transfus, NL-2333 ZC Leiden, Netherlands; [Cooper, Megan A.] Washington Univ, Sch Med, Dept Pediat, Div Rheumatol, St Louis, MO 63110 USA	Fehniger, TA (reprint author), Washington Univ, Sch Med, Dept Med, Div Oncol, St Louis, MO 63110 USA.	tfehnige@wustl.edutfehnige@wustl.edu	Rosario, Max/P-3111-2017; rosario, maxim/O-6098-2019		American Society of Hematology Foundation; Conquer Cancer Foundation of the American Society of Clinical Oncology; WUSM Siteman Cancer Center Developmental Research Award and Team Science Award; WUSM Institute of Clinical and Translational Research Award; Leukemia Specialized Program of Research Excellence Development Research Award [P50 CA171963]; Howard Hughes Medical Institute Medical Fellow Award; Translational TL1 program, NIH/National Cancer Institute (NCI) [F32 CA200253]; V Foundation for Cancer Research; Gabrielle's Angel Foundation for Cancer Research; Immunomonitoring Laboratory; Center for Human Immunology and Immunotherapy Programs; Biological Therapy Core; Small Animal Cancer Imaging Core; NCI Cancer Center Support grantUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [P30CA91842];  [P50 CA94056]	This work was supported by the American Society of Hematology Foundation, the Conquer Cancer Foundation of the American Society of Clinical Oncology, the WUSM Siteman Cancer Center Developmental Research Award and Team Science Award, the WUSM Institute of Clinical and Translational Research Award, the Leukemia Specialized Program of Research Excellence (P50 CA171963) Development Research Award, the Howard Hughes Medical Institute Medical Fellow Award, the Translational TL1 program, NIH/National Cancer Institute (NCI) grant F32 CA200253, the V Foundation for Cancer Research, and the Gabrielle's Angel Foundation for Cancer Research. Technical support was provided by the Immunomonitoring Laboratory (also supported by the Center for Human Immunology and Immunotherapy Programs), the Biological Therapy Core, and the Small Animal Cancer Imaging Core (also supported by P50 CA94056), which are supported by the NCI Cancer Center Support grant P30CA91842. We acknowledge the use of the Protein Production and Purification Facility for CyTOF mAb conjugation (P30 AR048335).	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Transl. Med.	SEP 21	2016	8	357							357ra123	10.1126/scitranslmed.aaf2341			12	Cell Biology; Medicine, Research & Experimental	Cell Biology; Research & Experimental Medicine	EE2UJ	WOS:000389440100003	27655849	Green Accepted, Bronze			2020-07-01	
J	Ornatsky, O; Bandura, D; Baranov, V; Nitz, M; Winnik, MA; Tanner, S				Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Nitz, Mark; Winnik, Mitchell A.; Tanner, Scott			Highly multiparametric analysis by mass cytometry	JOURNAL OF IMMUNOLOGICAL METHODS			English	Review						Mass cytometry; Metal-tagged antibodies; Metal-encoded beads; Multiparametric single cell assay; Flow cytometry; Bead array	ELEMENT-TAGGED IMMUNOASSAY; ACUTE MYELOID-LEUKEMIA; FLOW-CYTOMETRY; MULTIPLEX BIOASSAY; CELL SUBSETS; ARRAY ASSAYS; ICP-MS; BIOMARKERS; DIAGNOSIS; PANEL	This review paper describes a new technology, mass cytometry, that addresses applications typically run by flow cytometer analyzers, but extends the capability to highly multiparametric analysis. The detection technology is based on atomic mass spectrometry. It offers quantitation, specificity and dynamic range of mass spectrometry in a format that is familiar to flow cytometry practitioners. The mass cytometer does not require compensation, allowing the application of statistical techniques: this has been impossible given the constraints of fluorescence noise with traditional cytometry instruments. Instead of "colors" the mass cytometer "reads" the stable isotope tags attached to antibodies using metal-chelating labeling reagents. Because there are many available stable isotopes, and the mass spectrometer provides exquisite resolution between detection channels, many parameters can be measured as easily as one. For example, in a single tube the technique allows for the ready detection and characterization of the major cell subsets in blood or bone marrow. Here we describe mass cytometric immunophenotyping of human leukemia cell lines and leukemia patient samples, differential cell analysis of normal peripheral and umbilical cord blood: intracellular protein identification and metal-encoded bead arrays. (C) 2010 Elsevier B.V. All rights reserved.	[Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Nitz, Mark; Winnik, Mitchell A.; Tanner, Scott] Univ Toronto, Dept Chem, Toronto, ON M5S 3H6, Canada; [Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Tanner, Scott] DVS Sviences Inc, Richmond Hill, ON L4S 1Z5, Canada[Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Nitz, Mark; Winnik, Mitchell A.; Tanner, Scott] Univ Toronto, Dept Chem, Toronto, ON M5S 3H6, Canada; [Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Tanner, Scott] DVS Sviences Inc, Richmond Hill, ON L4S 1Z5, Canada	Ornatsky, O (reprint author), Univ Toronto, Dept Chem, 80 St George St, Toronto, ON M5S 3H6, Canada.	olga.ornatsky@utoronto.caolga.ornatsky@utoronto.ca	Winnik, Mitchell A/R-8621-2018		Genome Canada through the Ontario Genomics InstituteGenome Canada; Ontario Ministry of Research and InnovationMinistry of Research and Innovation, Ontario; National Institutes of Health [NIH]United States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [R01-GM076127]; NSERC CanadaNatural Sciences and Engineering Research Council of Canada; DVS Sciences Inc.	This project was funded by Genome Canada through the Ontario Genomics Institute, the Ontario Ministry of Research and Innovation, the National Institutes of Health [NIH grant R01-GM076127], NSERC Canada, and DVS Sciences Inc. Special thanks is extended to Dr. Qing Chang, Ontario Cancer Institute/Princess Margaret Hospital, University of Toronto, Ontario for the Panc-1 cells, and Dr. M. Milavsky and Dr. J. Wang, University Health Network (Toronto General Research Institute) for HSC and leukemia samples, respectively. Provision of leukemia bone marrow samples by the Quebec Leukemia Cell Bank (BCLQ) is gratefully acknowledged.	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Immunol. Methods	SEP 30	2010	361	1-2					1	20		10.1016/j.jim.2010.07.002			20	Biochemical Research Methods; Immunology	Biochemistry & Molecular Biology; Immunology	676FQ	WOS:000283896800001	20655312				2020-07-01	
J	Becher, B; Schlitzer, A; Chen, JM; Mair, F; Sumatoh, HR; Teng, KWW; Low, D; Ruedl, C; Riccardi-Castagnoli, P; Poidinger, M; Greter, M; Ginhoux, F; Newell, EW				Becher, Burkhard; Schlitzer, Andreas; Chen, Jinmiao; Mair, Florian; Sumatoh, Hermi R.; Teng, Karen Wei Weng; Low, Donovan; Ruedl, Christiane; Riccardi-Castagnoli, Paola; Poidinger, Michael; Greter, Melanie; Ginhoux, Florent; Newell, Evan W.			High-dimensional analysis of the murine myeloid cell system	NATURE IMMUNOLOGY			English	Article							CD11B(+) DENDRITIC CELLS; SUPPRESSOR-CELLS; STEADY-STATE; T-CELLS; GM-CSF; MONOCYTES; MACROPHAGES; IMMUNE; MOUSE; EXPRESSION	Advances in cell-fate mapping have revealed the complexity in phenotype, ontogeny and tissue distribution of the mammalian myeloid system. To capture this phenotypic diversity, we developed a 38-antibody panel for mass cytometry and used dimensionality reduction with machine learning-aided cluster analysis to build a composite of murine (mouse) myeloid cells in the steady state across lymphoid and nonlymphoid tissues. In addition to identifying all previously described myeloid populations, higher-order analysis allowed objective delineation of otherwise ambiguous subsets, including monocyte-macrophage intermediates and an array of granulocyte variants. Using mice that cannot sense granulocyte macrophage-colony stimulating factor GM-CSF (Csf2rb(-/-)), which have discrete alterations in myeloid development, we confirmed differences in barrier tissue dendritic cells, lung macrophages and eosinophils. The methodology further identified variations in the monocyte and innate lymphoid cell compartment that were unexpected, which confirmed that this approach is a powerful tool for unambiguous and unbiased characterization of the myeloid system.	[Becher, Burkhard; Schlitzer, Andreas; Chen, Jinmiao; Sumatoh, Hermi R.; Teng, Karen Wei Weng; Low, Donovan; Riccardi-Castagnoli, Paola; Poidinger, Michael; Ginhoux, Florent; Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Mair, Florian; Greter, Melanie] Univ Zurich, Inst Expt Immunol, CH-8091 Zurich, Switzerland; [Ruedl, Christiane] Nanyang Technol Univ, Sch Biol Sci, Singapore 639798, Singapore[Becher, Burkhard; Schlitzer, Andreas; Chen, Jinmiao; Sumatoh, Hermi R.; Teng, Karen Wei Weng; Low, Donovan; Riccardi-Castagnoli, Paola; Poidinger, Michael; Ginhoux, Florent; Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Mair, Florian; Greter, Melanie] Univ Zurich, Inst Expt Immunol, CH-8091 Zurich, Switzerland; [Ruedl, Christiane] Nanyang Technol Univ, Sch Biol Sci, Singapore 639798, Singapore	Newell, EW (reprint author), Univ Zurich, Inst Expt Immunol, Sternwartstr 2, CH-8091 Zurich, Switzerland.	becher@immunology.uzh.ch; evan_newell@immunol.a-staredu.sgbecher@immunology.uzh.ch; evan_newell@immunol.a-staredu.sg	Newell, Evan/AAE-9470-2020; Ruedl, christiane/A-2219-2011; Schlitzer, Andreas/R-2237-2016; Newell, Evan W/F-9711-2012	Newell, Evan/0000-0002-2889-243X; Ruedl, christiane/0000-0002-5599-6541; Schlitzer, Andreas/0000-0001-7662-3712; Newell, Evan W/0000-0002-2889-243X; Mair, Florian/0000-0001-6732-5449; Greter, Melanie/0000-0002-7220-5369; Poidinger, Michael/0000-0002-1047-2277; Becher, Burkhard/0000-0002-1541-7867	A*STAR/SIgNAgency for Science Technology & Research (ASTAR); Swiss National Science FoundationSwiss National Science Foundation (SNSF) [PP03P3_144781, 316030_150768, 310030_146130, CRSII3_136203]; European Union FP7 project TargetBraIn; University Research Priority Project 'Translational Cancer Research'; European Union FP7 project NeuroKine; European Union FP7 project Advanced T-cell Engineered for Cancer Therapy (ATECT)	The authors thank the SIgN community, the SIgN Flow Cytometry Facility and members of E.W.N.'s, F.G.'s and B.B.'s labs, and L.G. Ng for helpful discussion. Some antibodies used for generating the mass cytometry panels were provided by R. Balderas and A. Tiong (Becton Dickinson). The antibodies generated by BioXcell were provided for this analysis free of charge. B.B. performed this work while on sabbatical at A*STAR/SIgN. Supported by A*STAR/SIgN (P.R.-C., F.G., M.P., E.W.N.) and the Swiss National Science Foundation (PP03P3_144781 (M.G.), 316030_150768, 310030_146130 and CRSII3_136203 (B.B.)), European Union FP7 project TargetBraIn, NeuroKine, Advanced T-cell Engineered for Cancer Therapy (ATECT) and the University Research Priority Project 'Translational Cancer Research' (B.B.).	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Immunol.	DEC	2014	15	12					1181	1189		10.1038/ni.3006			9	Immunology	Immunology	AT3NR	WOS:000344841900015	25306126				2020-07-01	
J	Gury-BenAri, M; Thaiss, CA; Serafini, N; Winter, DR; Giladi, A; Lara-Astiaso, D; Levy, M; Salame, TM; Weiner, A; David, E; Shapiro, H; Dori-Bachash, M; Pevsner-Fischer, M; Lorenzo-Vivas, E; Keren-Shaul, H; Paul, F; Harmelin, A; Eberl, G; Itzkovitz, S; Tanay, A; Di Santo, JP; Elinav, E; Amit, I				Gury-BenAri, Meital; Thaiss, Christoph A.; Serafini, Nicolas; Winter, Deborah R.; Giladi, Amir; Lara-Astiaso, David; Levy, Maayan; Salame, Tomer Meir; Weiner, Assaf; David, Eyal; Shapiro, Hagit; Dori-Bachash, Mally; Pevsner-Fischer, Meirav; Lorenzo-Vivas, Erika; Keren-Shaul, Hadas; Paul, Franziska; Harmelin, Alon; Eberl, Gerard; Itzkovitz, Shalev; Tanay, Amos; Di Santo, James P.; Elinav, Eran; Amit, Ido			The Spectrum and Regulatory Landscape of Intestinal Innate Lymphoid Cells Are Shaped by the Microbiome	CELL			English	Article							TRANSCRIPTION FACTOR GATA3; ROR-GAMMA-T; RNA-SEQ; FUNCTIONAL PLASTICITY; COMMENSAL MICROFLORA; NKP46(+) CELLS; INFLAMMATION; HOMEOSTASIS; FATE; DIFFERENTIATION	Innate lymphoid cells (ILCs) are critical modulators of mucosal immunity, inflammation, and tissue homeostasis, but their full spectrum of cellular states and regulatory landscapes remains elusive. Here, we combine genome-wide RNA-seq, ChIP-seq, and ATAC-seq to compare the transcriptional and epigenetic identity of small intestinal ILCs, identifying thousands of distinct gene profiles and regulatory elements. Single-cell RNA-seq and flow and mass cytometry analyses reveal compartmentalization of cytokine expression and metabolic activity within the three classical ILC subtypes and highlight transcriptional states beyond the current canonical classification. In addition, using antibiotic intervention and germ-free mice, we characterize the effect of the microbiome on the ILC regulatory landscape and determine the response of ILCs to microbial colonization at the single-cell level. Together, our work characterizes the spectrum of transcriptional identities of small intestinal ILCs and describes how ILCs differentially integrate signals from the microbial microenvironment to generate phenotypic and functional plasticity.	[Gury-BenAri, Meital; Thaiss, Christoph A.; Winter, Deborah R.; Giladi, Amir; Lara-Astiaso, David; Levy, Maayan; Weiner, Assaf; David, Eyal; Shapiro, Hagit; Dori-Bachash, Mally; Pevsner-Fischer, Meirav; Lorenzo-Vivas, Erika; Keren-Shaul, Hadas; Paul, Franziska; Elinav, Eran; Amit, Ido] Weizmann Inst Sci, Dept Immunol, IL-76100 Rehovot, Israel; [Serafini, Nicolas; Di Santo, James P.] Inst Pasteur, Innate Immun Unit, F-75015 Paris, France; [Serafini, Nicolas; Di Santo, James P.] INSERM, U1223, Paris, France; [Salame, Tomer Meir] Weizmann Inst Sci, Biol Serv Unit, IL-76100 Rehovot, Israel; [Harmelin, Alon] Weizmann Inst Sci, Dept Vet Resources, IL-76100 Rehovot, Israel; [Eberl, Gerard] Inst Pasteur, Dept Immunol, F-75015 Paris, France; [Eberl, Gerard] CNRS, URA 1961, F-75015 Paris, France; [Itzkovitz, Shalev] Weizmann Inst Sci, Dept Mol Cell Biol, IL-76100 Rehovot, Israel; [Tanay, Amos] Weizmann Inst Sci, Dept Comp Sci & Appl Math, IL-76100 Rehovot, Israel; [Tanay, Amos] Weizmann Inst Sci, Dept Regulat Biol, IL-76100 Rehovot, Israel[Gury-BenAri, Meital; Thaiss, Christoph A.; Winter, Deborah R.; Giladi, Amir; Lara-Astiaso, David; Levy, Maayan; Weiner, Assaf; David, Eyal; Shapiro, Hagit; Dori-Bachash, Mally; Pevsner-Fischer, Meirav; Lorenzo-Vivas, Erika; Keren-Shaul, Hadas; Paul, Franziska; Elinav, Eran; Amit, Ido] Weizmann Inst Sci, Dept Immunol, IL-76100 Rehovot, Israel; [Serafini, Nicolas; Di Santo, James P.] Inst Pasteur, Innate Immun Unit, F-75015 Paris, France; [Serafini, Nicolas; Di Santo, James P.] INSERM, U1223, Paris, France; [Salame, Tomer Meir] Weizmann Inst Sci, Biol Serv Unit, IL-76100 Rehovot, Israel; [Harmelin, Alon] Weizmann Inst Sci, Dept Vet Resources, IL-76100 Rehovot, Israel; [Eberl, Gerard] Inst Pasteur, Dept Immunol, F-75015 Paris, France; [Eberl, Gerard] CNRS, URA 1961, F-75015 Paris, France; [Itzkovitz, Shalev] Weizmann Inst Sci, Dept Mol Cell Biol, IL-76100 Rehovot, Israel; [Tanay, Amos] Weizmann Inst Sci, Dept Comp Sci & Appl Math, IL-76100 Rehovot, Israel; [Tanay, Amos] Weizmann Inst Sci, Dept Regulat Biol, IL-76100 Rehovot, Israel	Elinav, E; Amit, I (reprint author), Weizmann Inst Sci, Dept Immunol, IL-76100 Rehovot, Israel.	eran.elinav@weizmann.ac.il; ido.amit@weizmann.ac.ileran.elinav@weizmann.ac.il; ido.amit@weizmann.ac.il	Di Santo, James/N-3143-2019; Di Santo, James P/M-4298-2014; SERAFINI, nicolas/S-5675-2019; Eberl, Gerard/A-1160-2013	Di Santo, James/0000-0002-7146-1862; Di Santo, James P/0000-0002-7146-1862; SERAFINI, nicolas/0000-0002-7440-4353; Eberl, Gerard/0000-0002-1119-5638; Lorenzo Vivas, Erika/0000-0002-8150-5902; Winter, Deborah/0000-0003-1806-673X; Paul, Franziska/0000-0003-0344-6764; Elinav, Eran/0000-0002-5775-2110	Boehringer Ingelheim Fonds PhD FellowshipBoehringer Ingelheim; Abisch Frenkel Foundation for the Promotion of Life Sciences; Gurwin Family Fund for Scientific Research; Leona M. and Harry B. Helmsley Charitable Trust; Crown Endowment Fund for Immunological Research; estate of J. Gitlitz; estate of L. Hershkovich; Benoziyo Endowment Fund for the Advancement of Science; Adelis Foundation; French National Center for Scientific Research (CNRS)Centre National de la Recherche Scientifique (CNRS); European Research CouncilEuropean Research Council (ERC); Marie Curie Integration grant; German-Israeli Foundation for Scientific Research and DevelopmentGerman-Israeli Foundation for Scientific Research and Development; Israel Science FoundationIsrael Science Foundation; Minerva Foundation; Rising Tide Foundation; Helmholtz FoundationHelmholtz Association; European Foundation for the Study of Diabetes; European Research CouncilEuropean Research Council (ERC) [309788]; Israel Science foundationIsrael Science Foundation [782/11]; BLUEPRINT FP7 consortium; Ernest and Bonnie Beutler Research Program of Excellence in Genomic Medicine; Minerva Stiftung research grant; National Human Genome Research Institute Center for Excellence in Genome Science [1P50HG006193]; Israeli Ministry of Science, Technology and Space; David and Fela Shapell Family Foundation; Abramson Family Center for Young Scientists	We thank the members of I.A.'s and E.E.'s labs for fruitful discussions, Carmit Bar-Nathan for GF mouse care taking, Elena Kartvelishvily for help with electron microscopy, Melanie Flach for technical advice, and Maayan Wigelmann for art work. C.A.T. received a Boehringer Ingelheim Fonds PhD Fellowship. E.E. is supported by Y. and R. Ungar; the Abisch Frenkel Foundation for the Promotion of Life Sciences; the Gurwin Family Fund for Scientific Research; the Leona M. and Harry B. Helmsley Charitable Trust; the Crown Endowment Fund for Immunological Research; the estate of J. Gitlitz; the estate of L. Hershkovich; the Benoziyo Endowment Fund for the Advancement of Science; the Adelis Foundation; J.L. and V. Schwartz; A. and G. Markovitz; A. and C. Adelson; the French National Center for Scientific Research (CNRS); D.L. Schwarz; the V.R. Schwartz Research Fellow Chair; L. Steinberg; J.N. Halpern; A. Edelheit, and by grants funded by the European Research Council; a Marie Curie Integration grant; the German-Israeli Foundation for Scientific Research and Development; the Israel Science Foundation; the Minerva Foundation; the Rising Tide Foundation; the Helmholtz Foundation; and the European Foundation for the Study of Diabetes. E.E. is the incumbent of the Rina Gudinski Career Development Chair and a senior fellow of the Canadian Institute For Advanced Research (CIFAR). I. A. is supported by the European Research Council (309788), the I-CORE for chromatin and RNA regulation, and personal grants from the Israel Science foundation (782/11) and the BLUEPRINT FP7 consortium, the Ernest and Bonnie Beutler Research Program of Excellence in Genomic Medicine, a Minerva Stiftung research grant, the National Human Genome Research Institute Center for Excellence in Genome Science (1P50HG006193), the Israeli Ministry of Science, Technology and Space, the David and Fela Shapell Family Foundation, and the Abramson Family Center for Young Scientists. I.A. is the incumbent of the Alan and Laraine Fischer Career Development Chair.	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J	Jiao, SP; Xia, WY; Yamaguchi, H; Wei, YK; Chen, MK; Hsu, JM; Hsu, JL; Yu, WH; Du, Y; Lee, HH; Li, CW; Chou, CK; Lim, SO; Chang, SS; Litton, J; Arun, B; Hortobagyi, GN; Hung, MC				Jiao, Shiping; Xia, Weiya; Yamaguchi, Hirohito; Wei, Yongkun; Chen, Mei-Kuang; Hsu, Jung-Mao; Hsu, Jennifer L.; Yu, Wen-Hsuan; Du, Yi; Lee, Heng-Huan; Li, Chia-Wei; Chou, Chao-Kai; Lim, Seung-Oe; Chang, Shih-Shin; Litton, Jennifer; Arun, Banu; Hortobagyi, Gabriel N.; Hung, Mien-Chie			PARP Inhibitor Upregulates PD-L1 Expression and Enhances Cancer-Associated Immunosuppression	CLINICAL CANCER RESEARCH			English	Article							CELL LUNG-CANCER; OVARIAN-CANCER; BREAST-CANCER; CHEMOTHERAPY; THERAPY; OVEREXPRESSION; PEMBROLIZUMAB; COMBINATION; ACTIVATION; PROVIDES	Purpose: To explore whether a cross-talk exists between PARP inhibition and PD-L1/PD-1 immune checkpoint axis, and determine whether blockade of PD-L1/PD-1 potentiates PARP inhibitor (PARPi) in tumor suppression. Experimental Design: Breast cancer cell lines, xenograft tumors, and syngeneic tumors treated with PARPi were assessed for PD-L1 expression by immunoblotting, IHC, and FACS analyses. The phospho-kinase antibody array screen was used to explore the underlying mechanism of PARPi-induced PD-L1 upregulation. The therapeutic efficacy of PARPi alone, PD-L1 blockade alone, or their combination was tested in a syngeneic tumor model. The tumor-infiltrating lymphocytes and tumor cells isolated from syngeneic tumors were analyzed by CyTOF and FACS to evaluate the activity of antitumor immunity in the tumor microenvironment. Results: PARPi upregulated PD-L1 expression in breast cancer cell lines and animal models. Mechanistically, PARPi inactivated GSK3b, which in turn enhanced PARPi-mediated PD-L1 upregulation. PARPi attenuated anticancer immunity via upregulation of PD-L1, and blockade of PD-L1 resensitized PARPi-treated cancer cells to T-cell killing. The combination of PARPi and anti-PD-L1 therapy compared with each agent alone significantly increased the therapeutic efficacy in vivo. Conclusions: Our study demonstrates a cross-talk between PARPi and tumor-associated immunosuppression and provides evidence to support the combination of PARPi and PD-L1 or PD-1 immune checkpoint blockade as a potential therapeutic approach to treat breast cancer. (C) 2017 AACR.	[Jiao, Shiping; Xia, Weiya; Yamaguchi, Hirohito; Wei, Yongkun; Chen, Mei-Kuang; Hsu, Jung-Mao; Hsu, Jennifer L.; Yu, Wen-Hsuan; Du, Yi; Lee, Heng-Huan; Li, Chia-Wei; Chou, Chao-Kai; Lim, Seung-Oe; Chang, Shih-Shin; Hung, Mien-Chie] Univ Texas MD Anderson Canc Ctr, Dept Mol & Cellular Oncol, 1515 Holcombe Blvd,Unit 108, Houston, TX 77030 USA; [Jiao, Shiping; Chen, Mei-Kuang; Yu, Wen-Hsuan; Hung, Mien-Chie] Univ Texas Houston, Grad Sch Biomed Sci, Houston, TX USA; [Hsu, Jennifer L.; Hung, Mien-Chie] China Med Univ, Grad Inst Biomed Sci, Taichung, Taiwan; [Hsu, Jennifer L.; Hung, Mien-Chie] China Med Univ, Ctr Mol Med, Taichung, Taiwan; [Hsu, Jennifer L.; Hung, Mien-Chie] Asia Univ, Dept Biotechnol, Taichung, Taiwan; [Litton, Jennifer; Arun, Banu; Hortobagyi, Gabriel N.] Univ Texas MD Anderson Canc Ctr, Dept Breast Med Oncol, Houston, TX 77030 USA[Jiao, Shiping; Xia, Weiya; Yamaguchi, Hirohito; Wei, Yongkun; Chen, Mei-Kuang; Hsu, Jung-Mao; Hsu, Jennifer L.; Yu, Wen-Hsuan; Du, Yi; Lee, Heng-Huan; Li, Chia-Wei; Chou, Chao-Kai; Lim, Seung-Oe; Chang, Shih-Shin; Hung, Mien-Chie] Univ Texas MD Anderson Canc Ctr, Dept Mol & Cellular Oncol, 1515 Holcombe Blvd,Unit 108, Houston, TX 77030 USA; [Jiao, Shiping; Chen, Mei-Kuang; Yu, Wen-Hsuan; Hung, Mien-Chie] Univ Texas Houston, Grad Sch Biomed Sci, Houston, TX USA; [Hsu, Jennifer L.; Hung, Mien-Chie] China Med Univ, Grad Inst Biomed Sci, Taichung, Taiwan; [Hsu, Jennifer L.; Hung, Mien-Chie] China Med Univ, Ctr Mol Med, Taichung, Taiwan; [Hsu, Jennifer L.; Hung, Mien-Chie] Asia Univ, Dept Biotechnol, Taichung, Taiwan; [Litton, Jennifer; Arun, Banu; Hortobagyi, Gabriel N.] Univ Texas MD Anderson Canc Ctr, Dept Breast Med Oncol, Houston, TX 77030 USA	Hung, MC (reprint author), Univ Texas MD Anderson Canc Ctr, Dept Mol & Cellular Oncol, 1515 Holcombe Blvd,Unit 108, Houston, TX 77030 USA.	mhung@mdanderson.orgmhung@mdanderson.org	Lim, Seung-Oe/D-3703-2011; Li, Chia-Wei/AAC-8660-2019; Hsu, Jung-Mao/AAC-9527-2019; Hsu, Jung-Mao/AAP-8045-2020; CHEN, MEI-KUANG/V-9284-2017	Li, Chia-Wei/0000-0002-2531-2866; CHEN, MEI-KUANG/0000-0001-8027-8990; Jiao, Shiping/0000-0001-6546-9369; Du, Yi/0000-0002-3969-2652; CHOU, CHAO-KAI/0000-0001-5231-5264	NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [CCSG CA016672]; Cancer Prevention & Research Institutes of Texas [DP150052, RP160710]; Breast Cancer Research Foundation; Patel Memorial Breast Cancer Endowment Fund; National Breast Cancer Foundation, Inc.; University of Texas MD Anderson Cancer Center-China Medical University and Hospital Sister Institution Fund; Ministry of Science and TechnologyMinistry of Education, Culture, Sports, Science and Technology, Japan (MEXT); International Research-intensive Centers of Excellence in Taiwan (I-RiCE) [MOST 105-2911-I-002-302]; Ministry of Health and Welfare, China Medical University Hospital Cancer Research Center of Excellence [MOHW106-TDU-B-212-144003]	This work was partially supported by the following: the NIH (CCSG CA016672), Cancer Prevention & Research Institutes of Texas (DP150052 and RP160710), Breast Cancer Research Foundation grant (to M.-C. Hung and G. N. Hortobagyi), Patel Memorial Breast Cancer Endowment Fund, National Breast Cancer Foundation, Inc., The University of Texas MD Anderson Cancer Center-China Medical University and Hospital Sister Institution Fund (to M.-C. Hung), Ministry of Science and Technology, International Research-intensive Centers of Excellence in Taiwan (I-RiCE; MOST 105-2911-I-002-302), and Ministry of Health and Welfare, China Medical University Hospital Cancer Research Center of Excellence (MOHW106-TDU-B-212-144003).	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Cancer Res.	JUL 15	2017	23	14					3711	3720		10.1158/1078-0432.CCR-16-3215			10	Oncology	Oncology	FA8EG	WOS:000405678400028	28167507	Bronze, Green Accepted			2020-07-01	
J	Wagner, A; Regev, A; Yosef, N				Wagner, Allon; Regev, Aviv; Yosef, Nir			Revealing the vectors of cellular identity with single-cell genomics	NATURE BIOTECHNOLOGY			English	Review							RNA-SEQ REVEALS; STOCHASTIC GENE-EXPRESSION; COPY-NUMBER VARIATION; MESSENGER-RNA; MASS CYTOMETRY; DIFFERENTIAL EXPRESSION; SEQUENCING DATA; CHROMATIN ACCESSIBILITY; TRANSCRIPTOMICS REVEALS; PARETO OPTIMALITY	Single-cell genomics has now made it possible to create a comprehensive atlas of human cells. At the same time, it has reopened definitions of a cell's identity and of the ways in which identity is regulated by the cell's molecular circuitry. Emerging computational analysis methods, especially in single-cell RNA sequencing (scRNA-seq), have already begun to reveal, in a data-driven way, the diverse simultaneous facets of a cell's identity, from discrete cell types to continuous dynamic transitions and spatial locations. These developments will eventually allow a cell to be represented as a superposition of ` basis vectors', each determining a different (but possibly dependent) aspect of cellular organization and function. However, computational methods must also overcome considerable challenges-from handling technical noise and data scale to forming new abstractions of biology. As the scale of single-cell experiments continues to increase, new computational approaches will be essential for constructing and characterizing a reference map of cell identities.	[Wagner, Allon; Yosef, Nir] Univ Calif Berkeley, Dept Elect Engn & Comp Sci, Berkeley, CA 94720 USA; [Wagner, Allon; Yosef, Nir] Univ Calif Berkeley, Ctr Computat Biol, Berkeley, CA USA; [Regev, Aviv] MIT, Dept Biol, Howard Hughes Med Inst, Cambridge, MA USA; [Regev, Aviv] Broad Inst MIT & Harvard, Cambridge, MA USA; [Yosef, Nir] MIT, Massachusetts Gen Hosp, Ragon Inst, Boston, MA USA; [Yosef, Nir] Harvard Univ, Boston, MA 02115 USA[Wagner, Allon; Yosef, Nir] Univ Calif Berkeley, Dept Elect Engn & Comp Sci, Berkeley, CA 94720 USA; [Wagner, Allon; Yosef, Nir] Univ Calif Berkeley, Ctr Computat Biol, Berkeley, CA USA; [Regev, Aviv] MIT, Dept Biol, Howard Hughes Med Inst, Cambridge, MA USA; [Regev, Aviv] Broad Inst MIT & Harvard, Cambridge, MA USA; [Yosef, Nir] MIT, Massachusetts Gen Hosp, Ragon Inst, Boston, MA USA; [Yosef, Nir] Harvard Univ, Boston, MA 02115 USA	Regev, A (reprint author), MIT, Dept Biol, Howard Hughes Med Inst, Cambridge, MA USA.; Regev, A (reprint author), Broad Inst MIT & Harvard, Cambridge, MA USA.	aregev@broadinstitute.orgaregev@broadinstitute.org	t, lf/AAJ-4924-2020	Wagner, Allon/0000-0001-5095-6009	US National Institute of Mental HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Mental Health (NIMH) [U01 MH105979]; Klarman Cell Observatory at the Broad Institute, NIH [P50 HG006193]; National Cancer InstituteUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [P30-CA14051]; NIH BRAIN grantUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [1U01MH105960-01]; NCIUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [1U24CA180922]; NIAIDUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [1U24AI118672-01]	We thank E. Lander, A.K. Shalek, R.B. Fletcher, O. Ram, and D. Stafford for helpful discussions, and L. Gaffney and A. Hupalowska for artwork. A.W. and N.Y. were supported in part by the BRAIN Initiative grant U01 MH105979 from the US National Institute of Mental Health. A.R. is an Investigator of the Howard Hughes Medical Institute and was supported by the Klarman Cell Observatory at the Broad Institute, NIH grant P50 HG006193, Koch Institute Support (core) grant P30-CA14051 from the National Cancer Institute, NIH BRAIN grant 1U01MH105960-01, NCI grant 1U24CA180922, and NIAID grant 1U24AI118672-01.	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Biotechnol.	NOV	2016	34	11					1145	1160		10.1038/nbt.3711			16	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	EB6PG	WOS:000387506500027	27824854	Green Accepted			2020-07-01	
J	Van Gassen, S; Callebaut, B; Van Helden, MJ; Lambrecht, BN; Demeester, P; Dhaene, T; Saeys, Y				Van Gassen, Sofie; Callebaut, Britt; Van Helden, Mary J.; Lambrecht, Bart N.; Demeester, Piet; Dhaene, Tom; Saeys, Yvan			FlowSOM: Using self-organizing maps for visualization and interpretation of cytometry data	CYTOMETRY PART A			English	Article						polychromatic flow cytometry; mass cytometry; exploratory data analysis; visualization method; self-organizing map; bioinformatics	IDENTIFICATION; ALGORITHM	The number of markers measured in both flow and mass cytometry keeps increasing steadily. Although this provides a wealth of information, it becomes infeasible to analyze these datasets manually. When using 2D scatter plots, the number of possible plots increases exponentially with the number of markers and therefore, relevant information that is present in the data might be missed. In this article, we introduce a new visualization technique, called FlowSOM, which analyzes Flow or mass cytometry data using a Self-Organizing Map. Using a two-level clustering and star charts, our algorithm helps to obtain a clear overview of how all markers are behaving on all cells, and to detect subsets that might be missed otherwise. R code is available at and will be made available at Bioconductor. (c) 2015 International Society for Advancement of Cytometry	[Van Gassen, Sofie; Callebaut, Britt; Demeester, Piet; Dhaene, Tom] Univ Ghent, Dept Informat Technol, IMinds, B-9000 Ghent, Belgium; [Van Gassen, Sofie; Van Helden, Mary J.; Lambrecht, Bart N.; Saeys, Yvan] VIB, Inflammat Res Ctr, Ghent, Belgium; [Van Gassen, Sofie; Van Helden, Mary J.; Lambrecht, Bart N.; Saeys, Yvan] Ghent Univ Hosp, Dept Resp Med, Ghent, Belgium[Van Gassen, Sofie; Callebaut, Britt; Demeester, Piet; Dhaene, Tom] Univ Ghent, Dept Informat Technol, IMinds, B-9000 Ghent, Belgium; [Van Gassen, Sofie; Van Helden, Mary J.; Lambrecht, Bart N.; Saeys, Yvan] VIB, Inflammat Res Ctr, Ghent, Belgium; [Van Gassen, Sofie; Van Helden, Mary J.; Lambrecht, Bart N.; Saeys, Yvan] Ghent Univ Hosp, Dept Resp Med, Ghent, Belgium	Van Gassen, S (reprint author), Univ Ghent VIB, Technol Pk 927, B-9052 Ghent, Zwijnaarde, Belgium.	sofie.vangassen@irc.vib-ugent.besofie.vangassen@irc.vib-ugent.be	Lambrecht, Bart N/K-2484-2014; Saeys, Yvan/C-1311-2009	Lambrecht, Bart N/0000-0003-4376-6834; Saeys, Yvan/0000-0002-0415-1506; Van Gassen, Sofie/0000-0002-7119-5330	Agency for Innovation by Science and Technology (IWT)Institute for the Promotion of Innovation by Science and Technology in Flanders (IWT); Ghent University Multidisciplinary Research Partnership Bioinformatics	Grant sponsors: Agency for Innovation by Science and Technology (IWT); Ghent University Multidisciplinary Research Partnership Bioinformatics	Aghaeepour N, 2013, NAT METHODS, V10, P228, DOI [10.1038/NMETH.2365, 10.1038/nmeth.2365]; Aghaeepour N, 2011, CYTOM PART A, V79A, P6, DOI 10.1002/cyto.a.21007; Amir ED, 2013, NAT BIOTECHNOL, V31, P545, DOI 10.1038/nbt.2594; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Cron A, 2013, PLOS COMPUT BIOL, V9, DOI 10.1371/journal.pcbi.1003130; Csardi G, 2006, INT J COMPLEX SYST, V1695, P1, DOI DOI 10.3724/SP.J.1087.2009.02191; Finak Greg, 2009, Advances in Bioinformatics, V2009, P247646, DOI 10.1155/2009/247646; KAMADA T, 1989, INFORM PROCESS LETT, V31, P7, DOI 10.1016/0020-0190(89)90102-6; KOHONEN T, 1990, P IEEE, V78, P1464, DOI 10.1109/5.58325; Lo K, 2008, CYTOM PART A, V73A, P321, DOI 10.1002/cyto.a.20531; Lo K, 2009, BMC BIOINFORMATICS, V10, DOI 10.1186/1471-2105-10-145; Naim I, 2014, CYTOM PART A, V85, P408, DOI 10.1002/cyto.a.22446; Pyne S, 2014, PLOS ONE, V9, DOI 10.1371/journal.pone.0100334; Pyne S, 2009, P NATL ACAD SCI USA, V106, P8519, DOI 10.1073/pnas.0903028106; Qiu P, 2011, NAT BIOTECHNOL, V29, P886, DOI 10.1038/nbt.1991; Wehrens R, 2007, J STAT SOFTW, V21, P1; WHITNEY VKM, 1972, COMMUN ACM, V15, P273, DOI 10.1145/361284.361299; Wilkerson M, 2013, CONSENSUSCLUSTERPLUS	18	180	180	5	25	WILEY	HOBOKEN	111 RIVER ST, HOBOKEN 07030-5774, NJ USA	1552-4922	1552-4930		CYTOM PART A	Cytom. Part A	JUL	2015	87A	7			SI		636	645		10.1002/cyto.a.22625			10	Biochemical Research Methods; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	CL3ZK	WOS:000356891200006	25573116				2020-07-01	
J	See, P; Dutertre, CA; Chen, JM; Gunther, P; McGovern, N; Irac, SE; Gunawan, M; Beyer, M; Handler, K; Duan, K; Bin Sumatoh, HR; Ruffin, N; Jouve, M; Gea-Mallorqui, E; Hennekam, RCM; Lim, T; Yip, CC; Wen, M; Malleret, B; Low, I; Shadan, NB; Fen, CFS; Tay, A; Lum, J; Zolezzi, F; Larbi, A; Poidinger, M; Chan, JKY; Chen, QF; Renia, L; Haniffa, M; Benaroch, P; Schlitzer, A; Schultze, JL; Newell, EW; Ginhoux, F				See, Peter; Dutertre, Charles-Antoine; Chen, Jinmiao; Gunther, Patrick; McGovern, Naomi; Irac, Sergio Erdal; Gunawan, Merry; Beyer, Marc; Handler, Kristian; Duan, Kaibo; Bin Sumatoh, Hermi Rizal; Ruffin, Nicolas; Jouve, Mabel; Gea-Mallorqui, Ester; Hennekam, Raoul C. M.; Lim, Tony; Yip, Chan Chung; Wen, Ming; Malleret, Benoit; Low, Ivy; Shadan, Nurhidaya Binte; Fen, Charlene Foong Shu; Tay, Alicia; Lum, Josephine; Zolezzi, Francesca; Larbi, Anis; Poidinger, Michael; Chan, Jerry K. Y.; Chen, Qingfeng; Renia, Laurent; Haniffa, Muzlifah; Benaroch, Philippe; Schlitzer, Andreas; Schultze, Joachim L.; Newell, Evan W.; Ginhoux, Florent			Mapping the human DC lineage through the integration of high-dimensional techniques	SCIENCE			English	Article							PLASMACYTOID DENDRITIC CELLS; INTERFERON-PRODUCING CELLS; INFLUENZA-VIRUS; T-CELLS; RNA-SEQ; EXPRESSION; PROGENITOR; SUBSETS; TRAJECTORIES; MACROPHAGES	Dendritic cells (DC) are professional antigen-presenting cells that orchestrate immune responses. The human DC population comprises two main functionally specialized lineages, whose origins and differentiation pathways remain incompletely defined. Here, we combine two high-dimensional technologies-single-cell messenger RNA sequencing (scmRNAseq) and cytometry by time-of-flight (CyTOF)-to identify human blood CD123(+)CD33(+)CD45RA(+)DC precursors (pre-DC). Pre-DC share surface markers with plasmacytoid DC (pDC) but have distinct functional properties that were previously attributed to pDC. Tracing the differentiation of DC from the bone marrow to the peripheral blood revealed that the pre-DC compartment contains distinct lineage-committed subpopulations, including one early uncommitted CD123(high) pre-DC subset and two CD45RA(+)CD123(low) lineage-committed subsets exhibiting functional differences. The discovery of multiple committed pre-DC populations opens promising new avenues for the therapeutic exploitation of DC subset-specific targeting.	[See, Peter; Dutertre, Charles-Antoine; Chen, Jinmiao; McGovern, Naomi; Duan, Kaibo; Bin Sumatoh, Hermi Rizal; Malleret, Benoit; Low, Ivy; Shadan, Nurhidaya Binte; Tay, Alicia; Lum, Josephine; Zolezzi, Francesca; Larbi, Anis; Poidinger, Michael; Chan, Jerry K. Y.; Renia, Laurent; Schlitzer, Andreas; Newell, Evan W.; Ginhoux, Florent] ASTAR, Singapore Immunol Network SIgN, 8A Biomed Grove,Immunos Bldg,Level 4, Singapore 138648, Singapore; [Dutertre, Charles-Antoine; Irac, Sergio Erdal; Wen, Ming] Duke NUS Med Sch, Program Emerging Infect Dis, 8 Coll Rd, Singapore 169857, Singapore; [Gunther, Patrick; Beyer, Marc; Handler, Kristian; Schultze, Joachim L.] Univ Bonn, Life & Med Sci LIMES Inst, Genom & Immunoregulat, D-32115 Bonn, Germany; [Gunawan, Merry; Haniffa, Muzlifah] Newcastle Univ, Inst Cellular Med, Newcastle Upon Tyne, Tyne & Wear, England; [Beyer, Marc; Schultze, Joachim L.] German Ctr Neurodegenerat Dis, Platform Single Cell Genom & Epigen, D-53175 Bonn, Germany; [Beyer, Marc; Schultze, Joachim L.] Univ Bonn, D-53175 Bonn, Germany; [Ruffin, Nicolas; Jouve, Mabel; Gea-Mallorqui, Ester; Benaroch, Philippe] PSL Res Univ, INSERM, U932, Inst Curie, F-75005 Paris, France; [Hennekam, Raoul C. M.] Univ Amsterdam, Acad Med Ctr, Dept Pediat, Amsterdam, Netherlands; [Lim, Tony] Singapore Gen Hosp, Dept Anat Pathol, Singapore, Singapore; [Yip, Chan Chung] Singapore Gen Hosp, Dept Hlth Promot Board HPB & Transplant Surg, Singapore, Singapore; [Malleret, Benoit] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Microbiol & Immunol, Singapore, Singapore; [Fen, Charlene Foong Shu] Singapore Hlth Serv Flow Cytometry Core Platform, 20 Coll Rd,Discovery Tower Level 10, Singapore 169856, Singapore; [Chan, Jerry K. Y.] KK Womens & Childrens Hosp, Div Obstet & Gynaecol, Dept Reprod Med, Singapore, Singapore; [Chan, Jerry K. Y.] Duke NUS Grad Med Sch, Canc & Stem Cell Biol Program, Singapore, Singapore; [Chan, Jerry K. Y.] Natl Univ Singapore, Yong Loo Lin Sch Med, Expt Fetal Med Grp, Singapore, Singapore; [Chen, Qingfeng] ASTAR, Humanized Mouse Unit, Inst Mol & Cell Biol, Singapore, Singapore; [Chen, Qingfeng] Univ Bonn, Myeloid Cell Biol Life & Med Sci LIMES Inst, D-53115 Bonn, Germany[See, Peter; Dutertre, Charles-Antoine; Chen, Jinmiao; McGovern, Naomi; Duan, Kaibo; Bin Sumatoh, Hermi Rizal; Malleret, Benoit; Low, Ivy; Shadan, Nurhidaya Binte; Tay, Alicia; Lum, Josephine; Zolezzi, Francesca; Larbi, Anis; Poidinger, Michael; Chan, Jerry K. Y.; Renia, Laurent; Schlitzer, Andreas; Newell, Evan W.; Ginhoux, Florent] ASTAR, Singapore Immunol Network SIgN, 8A Biomed Grove,Immunos Bldg,Level 4, Singapore 138648, Singapore; [Dutertre, Charles-Antoine; Irac, Sergio Erdal; Wen, Ming] Duke NUS Med Sch, Program Emerging Infect Dis, 8 Coll Rd, Singapore 169857, Singapore; [Gunther, Patrick; Beyer, Marc; Handler, Kristian; Schultze, Joachim L.] Univ Bonn, Life & Med Sci LIMES Inst, Genom & Immunoregulat, D-32115 Bonn, Germany; [Gunawan, Merry; Haniffa, Muzlifah] Newcastle Univ, Inst Cellular Med, Newcastle Upon Tyne, Tyne & Wear, England; [Beyer, Marc; Schultze, Joachim L.] German Ctr Neurodegenerat Dis, Platform Single Cell Genom & Epigen, D-53175 Bonn, Germany; [Beyer, Marc; Schultze, Joachim L.] Univ Bonn, D-53175 Bonn, Germany; [Ruffin, Nicolas; Jouve, Mabel; Gea-Mallorqui, Ester; Benaroch, Philippe] PSL Res Univ, INSERM, U932, Inst Curie, F-75005 Paris, France; [Hennekam, Raoul C. M.] Univ Amsterdam, Acad Med Ctr, Dept Pediat, Amsterdam, Netherlands; [Lim, Tony] Singapore Gen Hosp, Dept Anat Pathol, Singapore, Singapore; [Yip, Chan Chung] Singapore Gen Hosp, Dept Hlth Promot Board HPB & Transplant Surg, Singapore, Singapore; [Malleret, Benoit] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Microbiol & Immunol, Singapore, Singapore; [Fen, Charlene Foong Shu] Singapore Hlth Serv Flow Cytometry Core Platform, 20 Coll Rd,Discovery Tower Level 10, Singapore 169856, Singapore; [Chan, Jerry K. Y.] KK Womens & Childrens Hosp, Div Obstet & Gynaecol, Dept Reprod Med, Singapore, Singapore; [Chan, Jerry K. Y.] Duke NUS Grad Med Sch, Canc & Stem Cell Biol Program, Singapore, Singapore; [Chan, Jerry K. Y.] Natl Univ Singapore, Yong Loo Lin Sch Med, Expt Fetal Med Grp, Singapore, Singapore; [Chen, Qingfeng] ASTAR, Humanized Mouse Unit, Inst Mol & Cell Biol, Singapore, Singapore; [Chen, Qingfeng] Univ Bonn, Myeloid Cell Biol Life & Med Sci LIMES Inst, D-53115 Bonn, Germany	Ginhoux, F (reprint author), ASTAR, Singapore Immunol Network SIgN, 8A Biomed Grove,Immunos Bldg,Level 4, Singapore 138648, Singapore.	florent_ginhoux@immunol.a-star.edu.sgflorent_ginhoux@immunol.a-star.edu.sg	Malleret, Benoit/E-9159-2011; Schlitzer, Andreas/R-2237-2016; Irac, Sergio Erdal/G-6690-2014; Malleret, Benoit/AAK-4048-2020; Newell, Evan/AAE-9470-2020	Malleret, Benoit/0000-0001-9658-7528; Schlitzer, Andreas/0000-0001-7662-3712; Irac, Sergio Erdal/0000-0001-5622-1409; Malleret, Benoit/0000-0001-9658-7528; Newell, Evan/0000-0002-2889-243X; Beyer, Marc/0000-0001-9704-148X; Ruffin, Nicolas/0000-0002-3698-5505; Gunther, Patrick/0000-0002-9135-7819; McGovern, Naomi/0000-0001-5200-2698; Benaroch, Philippe/0000-0002-2655-0129; chen, jinmiao/0000-0001-7547-6423; Poidinger, Michael/0000-0002-1047-2277; Gea-Mallorqui, Ester/0000-0002-6915-074X; Dutertre, Charles-Antoine/0000-0001-7950-3652	Singapore Immunology Network core funding; Agency for Science, Technology and Research (A*STAR), SingaporeAgency for Science Technology & Research (ASTAR); A*STAR Graduate ScholarshipAgency for Science Technology & Research (ASTAR); Wellcome TrustWellcome Trust [WT 107931/Z/15/Z]; National Research Foundation SingaporeSingapore National Research Foundation [NMRC/BNIG/2026/2014]; French National Research AgencyFrench National Research Agency (ANR) [ANR-10-INSB-04, ANR-10-IDEX-0001-02 PSL, ANR-11-LABX-0043]; Agence Nationale de Recherche contre le SIDA et les hepatites virales (ANRS)ANRSFrench National Research Agency (ANR); Emmy-Noether fellowship of the German Research Foundation [SCHL 2116/1-1]; Biomedical Research Council SingaporeAgency for Science Technology & Research (ASTAR);  [Sonderforschungsbereich 645];  [704]	We thank L. Robinson of Insight Editing London for critical review and editing of the manuscript; P. Y. J. Ai from the SingHealth Flow Cytometry Core Platform; and M. L. Ng, S. H. Tan, and T. B. Lu from the Electron Microscope Unit of NUS for their assistance. This work was supported by Singapore Immunology Network core funding (F.G. and E.W.N.); Agency for Science, Technology and Research (A*STAR), Singapore; the A*STAR Graduate Scholarship (P. S.); the Wellcome Trust (WT 107931/Z/15/Z) (M. H.); and the National Research Foundation Singapore under its cooperative basic research grant new investigator grant (NMRC/BNIG/2026/2014) and administered by the Singapore Ministry of Health's National Medical Research Council (C.-A.D). This work was supported by the French National Research Agency through the "Investments for the Future" program (France-BioImaging, ANR-10-INSB-04), LABEX DCBIOL (ANR-10-IDEX-0001-02 PSL and ANR-11-LABX-0043), and grants from Agence Nationale de Recherche contre le SIDA et les hepatites virales (ANRS) (P.B., N.R., M.J., and E.G.-M.). J. L. S., M. B., and A. S. are members of the Excellence Cluster ImmunoSensation. J. L. S. is funded by Sonderforschungsbereich 645 and 704. A.S. is funded by an Emmy-Noether fellowship (SCHL 2116/1-1) of the German Research Foundation and a Young Investigator Award of the Biomedical Research Council Singapore. F.G. and P.S. are inventors on patent application 10201607246S held by A*STAR, which covers the methods for the identification, targeting, and isolation of human dendritic cell (DC) precursors ("preDC") and their use as biomarkers of inflammatory diseases. The MARS-seq, microfluidic scmRNAseq, and microarray data sets are deposited in the Gene Expression Omnibus under accession numbers GSE98052, GSE98011, and GSE80171, respectively.	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K., 2004, STAT APPL GENET MOL, V3, DOI DOI 10.2202/1544-6115.1027; Swiecki M, 2015, NAT REV IMMUNOL, V15, P471, DOI 10.1038/nri3865; Tenenbaum JB, 2000, SCIENCE, V290, P2319, DOI 10.1126/science.290.5500.2319; Trapnell C, 2014, NAT BIOTECHNOL, V32, P381, DOI 10.1038/nbt.2859; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; Yates A, 2016, NUCLEIC ACIDS RES, V44, pD710, DOI 10.1093/nar/gkv1157; Yu HS, 2015, PROTEIN CELL, V6, P297, DOI 10.1007/s13238-015-0140-x	61	179	181	10	72	AMER ASSOC ADVANCEMENT SCIENCE	WASHINGTON	1200 NEW YORK AVE, NW, WASHINGTON, DC 20005 USA	0036-8075	1095-9203		SCIENCE	Science	JUN 9	2017	356	6342			SI				eaag3009	10.1126/science.aag3009			11	Multidisciplinary Sciences	Science & Technology - Other Topics	EW9XC	WOS:000402871700045	28473638				2020-07-01	
J	Krieg, C; Nowicka, M; Guglietta, S; Schindler, S; Hartmann, FJ; Weber, LM; Dummer, R; Robinson, MD; Levesque, MP; Becher, B				Krieg, Carsten; Nowicka, Malgorzata; Guglietta, Silvia; Schindler, Sabrina; Hartmann, Felix J.; Weber, Lukas M.; Dummer, Reinhard; Robinson, Mark D.; Levesque, Mitchell P.; Becher, Burkhard			High-dimensional single-cell analysis predicts response to anti-PD-1 immunotherapy	NATURE MEDICINE			English	Article							DIFFERENTIAL EXPRESSION ANALYSIS; MASS CYTOMETRY; LUNG-CANCER; CLINICAL ACTIVITY; PERIPHERAL-BLOOD; PD-1 BLOCKADE; T-CELLS; MELANOMA; NIVOLUMAB; SAFETY	Immune-checkpoint blockade has revolutionized cancer therapy. In particular, inhibition of programmed cell death protein 1 (PD-1) has been found to be effective for the treatment of metastatic melanoma and other cancers. Despite a dramatic increase in progression-free survival, a large proportion of patients do not show durable responses. Therefore, predictive biomarkers of a clinical response are urgently needed. Here we used high-dimensional single-cell mass cytometry and a bioinformatics pipeline for the in-depth characterization of the immune cell subsets in the peripheral blood of patients with stage IV melanoma before and after 12 weeks of anti-PD-1 immunotherapy. During therapy, we observed a clear response to immunotherapy in the T cell compartment. However, before commencing therapy, a strong predictor of progression-free and overall survival in response to anti-PD-1 immunotherapy was the frequency of CD14(+)CD16-HLA-DRhi monocytes. We confirmed this by conventional flow cytometry in an independent, blinded validation cohort, and we propose that the frequency of monocytes in PBMCs may serve in clinical decision support.	[Krieg, Carsten; Hartmann, Felix J.; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, Zurich, Switzerland; [Nowicka, Malgorzata; Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Nowicka, Malgorzata; Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, SIB, Zurich, Switzerland; [Guglietta, Silvia] European Inst Oncol, Dept Expt Oncol, Milan, Italy; [Schindler, Sabrina; Dummer, Reinhard; Levesque, Mitchell P.] Univ Hosp Zurich, Dept Dermatol, Zurich, Switzerland; [Krieg, Carsten] Med Univ South Carolina, Dept Microbiol & Immunol, Hollings Canc Ctr, Charleston, SC 29425 USA; [Krieg, Carsten] Med Univ South Carolina, Dept Dermatol, Hollings Canc Ctr, Charleston, SC 29425 USA[Krieg, Carsten; Hartmann, Felix J.; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, Zurich, Switzerland; [Nowicka, Malgorzata; Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Nowicka, Malgorzata; Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, SIB, Zurich, Switzerland; [Guglietta, Silvia] European Inst Oncol, Dept Expt Oncol, Milan, Italy; [Schindler, Sabrina; Dummer, Reinhard; Levesque, Mitchell P.] Univ Hosp Zurich, Dept Dermatol, Zurich, Switzerland; [Krieg, Carsten] Med Univ South Carolina, Dept Microbiol & Immunol, Hollings Canc Ctr, Charleston, SC 29425 USA; [Krieg, Carsten] Med Univ South Carolina, Dept Dermatol, Hollings Canc Ctr, Charleston, SC 29425 USA	Krieg, C; Becher, B (reprint author), Univ Zurich, Inst Expt Immunol, Zurich, Switzerland.; Levesque, MP (reprint author), Univ Hosp Zurich, Dept Dermatol, Zurich, Switzerland.; Krieg, C (reprint author), Med Univ South Carolina, Dept Microbiol & Immunol, Hollings Canc Ctr, Charleston, SC 29425 USA.; Krieg, C (reprint author), Med Univ South Carolina, Dept Dermatol, Hollings Canc Ctr, Charleston, SC 29425 USA.	kriegc@musc.edu; mitchell.levesque@usz.ch; becher@immunology.uzh.chkriegc@musc.edu; mitchell.levesque@usz.ch; becher@immunology.uzh.ch	Robinson, Mark/O-2254-2019	Robinson, Mark/0000-0002-3048-5518; Hogan (Schindler), Sabrina/0000-0003-3994-8109; Becher, Burkhard/0000-0002-1541-7867; Hartmann, Felix/0000-0002-4174-2276; Weber, Lukas M./0000-0002-3282-1730; Krieg, Carsten/0000-0002-5145-7591	University Research Priority Program (URPP) in Translational Cancer Research; Swiss National Science FoundationSwiss National Science Foundation (SNSF) [310030_146130, 316030_150768]; European UnionEuropean Union (EU); European Training Network MELGEN	We thank V. Tosevski and T.M. Brodie (mass cytometry core facility, University of Zurich), A. Langer (Department of Dermatology, University of Zurich), and C. Beisel and K. Eschbach (Genomics Facility, ETH Basel) for excellent technical assistance and N. Nunes, B. Chatterjee, E. Terskikh, and C. Gujer (all from the Institute of Experimental Immunology, University Zurich), A. Zollinger (Swiss Institute of Bioinformatics, Lausanne), all members of the COST Action BM1404 Mye-EUNITER (http://www.mye-euniter.eu/), and P. Cheng (University of Zurich) for discussions. We also thank C. Guglietta for graphical design and layout. This work received funding from the University Research Priority Program (URPP) in Translational Cancer Research (C. K.), the Swiss National Science Foundation (grants 310030_146130 and 316030_150768; B. B.), the European Union FP7 project ATECT (B. B.), and the European Training Network MELGEN (M.P.L.).	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Med.	FEB	2018	24	2					144	+		10.1038/nm.4466			11	Biochemistry & Molecular Biology; Cell Biology; Medicine, Research & Experimental	Biochemistry & Molecular Biology; Cell Biology; Research & Experimental Medicine	FV1MQ	WOS:000424327200011	29309059	Green Accepted			2020-07-01	
J	Bruggner, RV; Bodenmiller, B; Dill, DL; Tibshirani, RJ; Nolan, GP				Bruggner, Robert V.; Bodenmiller, Bernd; Dill, David L.; Tibshirani, Robert J.; Nolan, Garry P.			Automated identification of stratifying signatures in cellular subpopulations	PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA			English	Article						informatics; biomarker discovery	FLOW-CYTOMETRY DATA; MASS CYTOMETRY; REGRESSION; RESPONSES; SELECTION; SUBSETS	Elucidation and examination of cellular subpopulations that display condition-specific behavior can play a critical contributory role in understanding disease mechanism, as well as provide a focal point for development of diagnostic criteria linking such a mechanism to clinical prognosis. Despite recent advancements in single-cell measurement technologies, the identification of relevant cell subsets through manual efforts remains standard practice. As new technologies such as mass cytometry increase the parameterization of single-cell measurements, the scalability and subjectivity inherent in manual analyses slows both analysis and progress. We therefore developed Citrus (cluster identification, characterization, and regression), a data-driven approach for the identification of stratifying subpopulations in multidimensional cytometry datasets. The methodology of Citrus is demonstrated through the identification of known and unexpected pathway responses in a dataset of stimulated peripheral blood mononuclear cells measured by mass cytometry. Additionally, the performance of Citrus is compared with that of existing methods through the analysis of several publicly available datasets. As the complexity of flow cytometry datasets continues to increase, methods such as Citrus will be needed to aid investigators in the performance of unbiased-and potentially more thorough-correlation-based mining and inspection of cell subsets nested within high-dimensional datasets.	[Bruggner, Robert V.] Stanford Univ, Sch Med, Biomed Informat Training Program, Stanford, CA 94305 USA; [Bruggner, Robert V.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Dill, David L.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Tibshirani, Robert J.] Stanford Univ, Dept Hlth Res & Policy, Stanford, CA 94305 USA; [Tibshirani, Robert J.] Stanford Univ, Dept Stat, Stanford, CA 94305 USA; [Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, CH-8057 Zurich, Switzerland[Bruggner, Robert V.] Stanford Univ, Sch Med, Biomed Informat Training Program, Stanford, CA 94305 USA; [Bruggner, Robert V.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Dill, David L.] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Tibshirani, Robert J.] Stanford Univ, Dept Hlth Res & Policy, Stanford, CA 94305 USA; [Tibshirani, Robert J.] Stanford Univ, Dept Stat, Stanford, CA 94305 USA; [Bodenmiller, Bernd] Univ Zurich, Inst Mol Life Sci, CH-8057 Zurich, Switzerland	Tibshirani, RJ (reprint author), Stanford Univ, Dept Hlth Res & Policy, Stanford, CA 94305 USA.	tibs@stanford.edu; gnolan@stanford.edutibs@stanford.edu; gnolan@stanford.edu	Nolan, Garry/AAE-7903-2019		National Library of Medicine Training Grant [T15 LM007033]; Swiss National Science FoundationSwiss National Science Foundation (SNSF); European Molecular Biology OrganizationEuropean Molecular Biology Organization (EMBO); Marie Curie International Outgoing FellowshipEuropean Union (EU); National Cancer Institute GrantUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [U54CA149145]; National Science FoundationNational Science Foundation (NSF) [DMS-9971405]; National Institutes of Health (NIH)United States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [N01-HV-28183]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U54CA149145, UL1RR025744, 0158 G KB065, 1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, 41000411217, 5 24927, P01 CA034233-22A1, PN2EY018228, RFA CA 09-009, RFA CA 09 011, U19 AI057229]; California Institute for Regenerative MedicineCalifornia Institute for Regenerative Medicine [DR1-01477, RB2-01592]; European CommissionEuropean Commission Joint Research Centre [HEALTH. 2010.1.2-1]; U.S. Food and Drug Administration [HHSF223201210194C: BAA-12-00118]; U.S. Department of DefenseUnited States Department of Defense [W81XWH-12-1-0591 OCRP-TIA NWC]	We thank the two manuscript referees for their many helpful comments and feedback. We also wish to thank N. Kotecha for additional feedback on the manuscript, R. Finck for insight during method development, and M. Linderman for his work on the Rclusterpp R package. R.V.B. is supported by National Library of Medicine Training Grant T15 LM007033. B.B. is supported by grants from the Swiss National Science Foundation, the European Molecular Biology Organization, and a Marie Curie International Outgoing Fellowship. D.L.D. is supported by National Cancer Institute Grant U54CA149145. R.J.T. is supported by National Science Foundation Grant DMS-9971405 and National Institutes of Health (NIH) Grant N01-HV-28183. G.P.N. is supported by NIH Grants UL1RR025744, 0158 G KB065, 1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, 41000411217, 5 24927, P01 CA034233-22A1, P01 CA034233-22A1, PN2EY018228, RFA CA 09-009, RFA CA 09 011, U19 AI057229, and U54CA149145; California Institute for Regenerative Medicine Grants DR1-01477 and RB2-01592; European Commission Grant HEALTH. 2010.1.2-1; U.S. Food and Drug Administration Grant HHSF223201210194C: BAA-12-00118; and U.S. Department of Defense Grant W81XWH-12-1-0591 OCRP-TIA NWC.	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Natl. Acad. Sci. U. S. A.	JUL 1	2014	111	26					E2770	E2777		10.1073/pnas.1408792111			8	Multidisciplinary Sciences	Science & Technology - Other Topics	AK0QG	WOS:000338118900020	24979804	Green Published, Bronze			2020-07-01	
J	Simoni, Y; Fehlings, M; Kloverpris, HN; McGovern, N; Koo, SL; Loh, CY; Lim, S; Kurioka, A; Fergusson, JR; Tang, CL; Kam, MH; Dennis, K; Lim, TKH; Fui, ACY; Hoong, CW; Chan, JKY; de Lafaille, MC; Narayanan, S; Baig, S; Shabeer, M; Toh, SAES; Tan, HKK; Anicete, R; Tan, EH; Takano, A; Klenerman, P; Leslie, A; Tan, DSW; Tan, IB; Ginhoux, F; Newell, EW				Simoni, Yannick; Fehlings, Michael; Kloverpris, Henrik N.; McGovern, Naomi; Koo, Si-Lin; Loh, Chiew Yee; Lim, Shawn; Kurioka, Ayako; Fergusson, Joannah R.; Tang, Choong-Leong; Kam, Ming Hian; Dennis, Koh; Lim, Tony Kiat Hon; Fui, Alexander Chung Yaw; Hoong, Chan Weng; Chan, Jerry Kok Yen; de Lafaille, Maria Curotto; Narayanan, Sriram; Baig, Sonia; Shabeer, Muhammad; Toh, Sue-Anne Ee Shiow; Tan, Henry Kun Kiaang; Anicete, Rosslyn; Tan, Eng-Huat; Takano, Angela; Klenerman, Paul; Leslie, Alasdair; Tan, Daniel S. W.; Tan, Iain Beehuat; Ginhoux, Florent; Newell, Evan W.			Human Innate Lymphoid Cell Subsets Possess Tissue-Type Based Heterogeneity in Phenotype and Frequency	IMMUNITY			English	Article							INFLAMMATION; EXPRESSION; INTERLEUKIN-2; HOMEOSTASIS; ACTIVATION; DISEASE; UNIQUE; IL-18; FATE	Animal models have highlighted the importance of innate lymphoid cells (ILCs) in multiple immune responses. However, technical limitations have hampered adequate characterization of ILCs in humans. Here, we used mass cytometry including a broad range of surface markers and transcription factors to accurately identify and profile ILCs across healthy and inflamed tissue types. High dimensional analysis allowed for clear phenotypic delineation of ILC2 and ILC3 subsets. We were not able to detect ILC1 cells in any of the tissues assessed, however, we identified intra-epithelial (ie) ILC1-like cells that represent a broader category of NK cells in mucosal and non-mucosal pathological tissues. In addition, we have revealed the expression of phenotypic molecules that have not been previously described for ILCs. Our analysis shows that human ILCs are highly heterogeneous cell types between individuals and tissues. It also provides a global, comprehensive, and detailed description of ILC heterogeneity in humans across patients and tissues.	[Simoni, Yannick; Fehlings, Michael; McGovern, Naomi; Loh, Chiew Yee; Lim, Shawn; de Lafaille, Maria Curotto; Ginhoux, Florent; Newell, Evan W.] Agcy Sci Technol & Res, Singapore Immunol Network SIgN, Singapore 138648, Singapore; [Kloverpris, Henrik N.; Leslie, Alasdair] Africa Hlth Res Inst, ZA-4001 Durban, South Africa; [Kloverpris, Henrik N.] Univ Copenhagen, Dept Immunol & Microbiol, DK-1165 Copenhagen, Denmark; [Koo, Si-Lin; Tan, Eng-Huat; Tan, Daniel S. W.; Tan, Iain Beehuat] Natl Canc Ctr Singapore, Div Med Oncol, Singapore 169610, Singapore; [Kurioka, Ayako; Fergusson, Joannah R.; Klenerman, Paul] Univ Oxford, Peter Medawar Bldg Pathogen Res, Oxford OX1 3SY, England; [Tang, Choong-Leong; Kam, Ming Hian; Dennis, Koh] Singapore Gen Hosp, Dept Colorectal Surg, Singapore 169856, Singapore; [Dennis, Koh] Mt Elizabeth Med Ctr, Colorectal Practice, Singapore 228510, Singapore; [Lim, Tony Kiat Hon; Takano, Angela] Singapore Gen Hosp, Dept Anat Pathol, Singapore 169608, Singapore; [Fui, Alexander Chung Yaw] Singapore Gen Hosp, Dept Hepatopancreatobiliary Transplant Surg, Singapore 169608, Singapore; [Hoong, Chan Weng] Singapore Gen Hosp, Dept Upper GI Bariatr Surg, Singapore 169608, Singapore; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Singapore 229899, Singapore; [Chan, Jerry Kok Yen; Toh, Sue-Anne Ee Shiow; Tan, Iain Beehuat] Duke NUS Grad Med Sch, Singapore 169857, Singapore; [Narayanan, Sriram] Agcy Sci Technol & Res, Inst Mol & Cell Biol, Singapore 138673, Singapore; [Baig, Sonia; Toh, Sue-Anne Ee Shiow] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Med, Singapore 119228, Singapore; [de Lafaille, Maria Curotto] NYU, Sch Med, 550 1st Ave, New York, NY 10012 USA; [Toh, Sue-Anne Ee Shiow] Natl Univ Hlth Syst, Dept Med, Singapore 119228, Singapore; [Toh, Sue-Anne Ee Shiow] Univ Penn, Perelman Sch Med, Philadelphia, PA 19104 USA; [Tan, Henry Kun Kiaang; Anicete, Rosslyn] KK Womens & Childrens Hosp, Dept Otolaryngol, Singapore 229899, Singapore; [Tan, Daniel S. W.; Tan, Iain Beehuat] Agcy Sci Technol & Res, Genome Inst Singapore, Singapore 138672, Singapore[Simoni, Yannick; Fehlings, Michael; McGovern, Naomi; Loh, Chiew Yee; Lim, Shawn; de Lafaille, Maria Curotto; Ginhoux, Florent; Newell, Evan W.] Agcy Sci Technol & Res, Singapore Immunol Network SIgN, Singapore 138648, Singapore; [Kloverpris, Henrik N.; Leslie, Alasdair] Africa Hlth Res Inst, ZA-4001 Durban, South Africa; [Kloverpris, Henrik N.] Univ Copenhagen, Dept Immunol & Microbiol, DK-1165 Copenhagen, Denmark; [Koo, Si-Lin; Tan, Eng-Huat; Tan, Daniel S. W.; Tan, Iain Beehuat] Natl Canc Ctr Singapore, Div Med Oncol, Singapore 169610, Singapore; [Kurioka, Ayako; Fergusson, Joannah R.; Klenerman, Paul] Univ Oxford, Peter Medawar Bldg Pathogen Res, Oxford OX1 3SY, England; [Tang, Choong-Leong; Kam, Ming Hian; Dennis, Koh] Singapore Gen Hosp, Dept Colorectal Surg, Singapore 169856, Singapore; [Dennis, Koh] Mt Elizabeth Med Ctr, Colorectal Practice, Singapore 228510, Singapore; [Lim, Tony Kiat Hon; Takano, Angela] Singapore Gen Hosp, Dept Anat Pathol, Singapore 169608, Singapore; [Fui, Alexander Chung Yaw] Singapore Gen Hosp, Dept Hepatopancreatobiliary Transplant Surg, Singapore 169608, Singapore; [Hoong, Chan Weng] Singapore Gen Hosp, Dept Upper GI Bariatr Surg, Singapore 169608, Singapore; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Singapore 229899, Singapore; [Chan, Jerry Kok Yen; Toh, Sue-Anne Ee Shiow; Tan, Iain Beehuat] Duke NUS Grad Med Sch, Singapore 169857, Singapore; [Narayanan, Sriram] Agcy Sci Technol & Res, Inst Mol & Cell Biol, Singapore 138673, Singapore; [Baig, Sonia; Toh, Sue-Anne Ee Shiow] Natl Univ Singapore, Yong Loo Lin Sch Med, Dept Med, Singapore 119228, Singapore; [de Lafaille, Maria Curotto] NYU, Sch Med, 550 1st Ave, New York, NY 10012 USA; [Toh, Sue-Anne Ee Shiow] Natl Univ Hlth Syst, Dept Med, Singapore 119228, Singapore; [Toh, Sue-Anne Ee Shiow] Univ Penn, Perelman Sch Med, Philadelphia, PA 19104 USA; [Tan, Henry Kun Kiaang; Anicete, Rosslyn] KK Womens & Childrens Hosp, Dept Otolaryngol, Singapore 229899, Singapore; [Tan, Daniel S. W.; Tan, Iain Beehuat] Agcy Sci Technol & Res, Genome Inst Singapore, Singapore 138672, Singapore	Simoni, Y (reprint author), Agcy Sci Technol & Res, Singapore Immunol Network SIgN, Singapore 138648, Singapore.	yannick_simoni@immunol.a-star.edu.sg; evan_newell@immunol.a-star.edu.sgyannick_simoni@immunol.a-star.edu.sg; evan_newell@immunol.a-star.edu.sg	Newell, Evan W/F-9711-2012; Newell, Evan/AAE-9470-2020	Newell, Evan W/0000-0002-2889-243X; Newell, Evan/0000-0002-2889-243X; leslie, alasdair/0000-0003-2538-6467; McGovern, Naomi/0000-0001-5200-2698; Koo, Si-Lin/0000-0002-8217-3321; Kloverpris, Henrik/0000-0002-2165-3313	A-STAR/SIgN core funding; A-STAR/SIgN immunomonitoring platform funding; Translational & Clinical Research grand from the National Medical Research Council, Singapore [NMRC/TCR/007-NCC/2013]; National Cancer Center Singapore [CG/007/2013-SD2 (NCCSPG-YR2015-JUL-13)]; NIHR Biomedical Research Centre, OxfordNational Institute for Health Research (NIHR); NIHR Senior Fellowship;  [WT109965MA]; Lundbeck FoundationLundbeckfonden [R151-2013-14624]; National Institute for Health ResearchNational Institute for Health Research (NIHR) [NF-SI-0515-10005]	The authors thank all members of E.N. and F.G. laboratory, Etienne Becht, Harsimran Singh, the SIgN community, and the SIgN flow cytometry and luminex facilities. This study was funded by A-STAR/SIgN core funding, A-STAR/SIgN immunomonitoring platform funding, a Translational & Clinical Research grand from the National Medical Research Council, Singapore (NMRC/TCR/007-NCC/2013) and the National Cancer Center Singapore CG/007/2013-SD2 (NCCSPG-YR2015-JUL-13). P.K. is supported by NIHR Biomedical Research Centre, Oxford, an NIHR Senior Fellowship and WT109965MA.	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J	Zunder, ER; Finck, R; Behbehani, GK; Amir, ED; Krishnaswamy, S; Gonzalez, VD; Lorang, CG; Bjornson, Z; Spitzer, MH; Bodenmiller, B; Fantl, WJ; Pe'er, D; Nolan, GP				Zunder, Eli R.; Finck, Rachel; Behbehani, Gregory K.; Amir, El-ad D.; Krishnaswamy, Smita; Gonzalez, Veronica D.; Lorang, Cynthia G.; Bjornson, Zach; Spitzer, Matthew H.; Bodenmiller, Bernd; Fantl, Wendy J.; Pe'er, Dana; Nolan, Garry P.			Palladium-based mass tag cell barcoding with a doublet-filtering scheme and single-cell deconvolution algorithm	NATURE PROTOCOLS			English	Article							CYTOMETRY DATA; SAMPLES	Mass-tag cell barcoding (MCB) labels individual cell samples with unique combinatorial barcodes, after which they are pooled for processing and measurement as a single multiplexed sample. The MCB method eliminates variability between samples in antibody staining and instrument sensitivity, reduces antibody consumption and shortens instrument measurement time. Here we present an optimized MCB protocol. The use of palladium-based labeling reagents expands the number of measurement channels available for mass cytometry and reduces interference with lanthanide-based antibody measurement. An error-detecting combinatorial barcoding scheme allows cell doublets to be identified and removed from the analysis. A debarcoding algorithm that is single cell-based rather than population-based improves the accuracy and efficiency of sample deconvolution. This debarcoding algorithm has been packaged into software that allows rapid and unbiased sample deconvolution. The MCB procedure takes 3-4 h, not including sample acquisition time of similar to 1 h per million cells.	[Zunder, Eli R.; Finck, Rachel; Behbehani, Gregory K.; Gonzalez, Veronica D.; Lorang, Cynthia G.; Bjornson, Zach; Spitzer, Matthew H.; Bodenmiller, Bernd; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Hematol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Oncol, Stanford, CA 94305 USA; [Amir, El-ad D.; Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA[Zunder, Eli R.; Finck, Rachel; Behbehani, Gregory K.; Gonzalez, Veronica D.; Lorang, Cynthia G.; Bjornson, Zach; Spitzer, Matthew H.; Bodenmiller, Bernd; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Hematol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Oncol, Stanford, CA 94305 USA; [Amir, El-ad D.; Krishnaswamy, Smita; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA	Nolan, GP (reprint author), Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Spitzer, Matthew/0000-0002-5291-3819; Pe'er, Dana/0000-0002-9259-8817	US National Institutes of Health (NIH)United States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, U54CA149145, N01-HV-00242, 1U19AI100627, 5R01AI07372405, R01CA184968, 1 R33 CA183654, NRSA F32 GM093508-01, 152175.5041015.0412, R01 A1073724]; NIH-the Baylor Research Institute [41000411217]; NIH-Northrop Grumman Corp. [7500108142]; California Institute for Regenerative Medicine (CIRM)California Institute for Regenerative Medicine [DR1-01477, RB2-01592]; US Department of DefenseUnited States Department of Defense [OC110674]; European CommissionEuropean Commission Joint Research Centre [Health.2010.1.2-1]; US Food and Drug Administration [HHSF223201210194C]; Bill and Melinda Gates FoundationGates Foundation [OPP 1017093]; Alliance for Lupus Research, the Lymphoma Research Foundation; Entertainment Industry Foundation (National Women's Cancer Research Alliance grant); National Science Foundation CAREER awardNational Science Foundation (NSF) [MCB-1149728]; Packard Fellowship for Science and Engineering; Rachford and Carlota A. Harris Endowed Professorship	We thank A. Trejo and A. Jager for mass cytometry quality control and instrument maintenance. We thank E. Simonds and P. Krutzik for their helpful discussions. This work was supported by grants from the US National Institutes of Health (NIH) (U19 AI057229, U54CA149145, N01-HV-00242, 1U19AI100627, 5R01AI07372405, R01CA184968, 1 R33 CA183654, NRSA F32 GM093508-01, 152175.5041015.0412 and R01 A1073724), NIH-the Baylor Research Institute (41000411217), the NIH-Northrop Grumman Corp. (7500108142), the California Institute for Regenerative Medicine (CIRM) (DR1-01477, RB2-01592), the US Department of Defense (OC110674), the European Commission (Health.2010.1.2-1), the US Food and Drug Administration (HHSF223201210194C), the Bill and Melinda Gates Foundation (OPP 1017093), the Alliance for Lupus Research, the Lymphoma Research Foundation, the Entertainment Industry Foundation (National Women's Cancer Research Alliance grant) and the National Science Foundation CAREER award (MCB-1149728). D.P. holds a Packard Fellowship for Science and Engineering. G.P.N. holds the Rachford and Carlota A. Harris Endowed Professorship.	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Protoc.	FEB	2015	10	2					316	333		10.1038/nprot.2015.020			18	Biochemical Research Methods	Biochemistry & Molecular Biology	CE6ET	WOS:000351930100007	25612231	Green Accepted			2020-07-01	
J	Swadling, L; Capone, S; Antrobus, RD; Brown, A; Richardson, R; Newell, EW; Halliday, J; Kelly, C; Bowen, D; Fergusson, J; Kurioka, A; Ammendola, V; Del Sorbo, M; Grazioli, F; Esposito, ML; Siani, L; Traboni, C; Hill, A; Colloca, S; Davis, M; Nicosia, A; Cortese, R; Folgori, A; Klenerman, P; Barnes, E				Swadling, Leo; Capone, Stefania; Antrobus, Richard D.; Brown, Anthony; Richardson, Rachel; Newell, Evan W.; Halliday, John; Kelly, Christabel; Bowen, Dan; Fergusson, Joannah; Kurioka, Ayako; Ammendola, Virginia; Del Sorbo, Mariarosaria; Grazioli, Fabiana; Esposito, Maria Luisa; Siani, Loredana; Traboni, Cinzia; Hill, Adrian; Colloca, Stefano; Davis, Mark; Nicosia, Alfredo; Cortese, Riccardo; Folgori, Antonella; Klenerman, Paul; Barnes, Eleanor			A human vaccine strategy based on chimpanzee adenoviral and MVA vectors that primes, boosts, and sustains functional HCV-specific T cell memory	SCIENCE TRANSLATIONAL MEDICINE			English	Article							HEPATITIS-C VIRUS; SINGLE-SOURCE OUTBREAK; IMMUNE-RESPONSES; VIRAL CLEARANCE; NEUTRALIZING ANTIBODIES; CYTOKINE EXPRESSION; ACUTE INFECTION; STAGE MALARIA; PERSISTENCE; CHALLENGE	A protective vaccine against hepatitis C virus (HCV) remains an unmet clinical need. HCV infects millions of people worldwide and is a leading cause of liver cirrhosis and hepatocellular cancer. Animal challenge experiments, immunogenetics studies, and assessment of host immunity during acute infection highlight the critical role that effective T cell immunity plays in viral control. In this first-in-man study, we have induced antiviral immunity with functional characteristics analogous to those associated with viral control in natural infection, and improved upon a vaccine based on adenoviral vectors alone. We assessed a heterologous prime-boost vaccination strategy based on a replicative defective simian adenoviral vector (ChAd3) and modified vaccinia Ankara (MVA) vector encoding the NS3, NS4, NS5A, and NS5B proteins of HCV genotype 1b. Analysis used single-cell mass cytometry and human leukocyte antigen class I peptide tetramer technology in healthy human volunteers. We show that HCV-specific T cells induced by ChAd3 are optimally boosted with MVA, and generate very high levels of both CD8(+) and CD4(+) HCV-specific T cells targeting multiple HCV antigens. Sustained memory and effector T cell populations are generated, and T cell memory evolved over time with improvement of quality (proliferation and polyfunctionality) after heterologous MVA boost. We have developed an HCV vaccine strategy, with durable, broad, sustained, and balanced T cell responses, characteristic of those associated with viral control, paving the way for the first efficacy studies of a prophylactic HCV vaccine.	[Swadling, Leo; Antrobus, Richard D.; Brown, Anthony; Richardson, Rachel; Halliday, John; Kelly, Christabel; Bowen, Dan; Fergusson, Joannah; Kurioka, Ayako; Hill, Adrian; Klenerman, Paul; Barnes, Eleanor] Univ Oxford, Nuffield Dept Med, Oxford OX1 3SY, England; [Capone, Stefania; Ammendola, Virginia; Del Sorbo, Mariarosaria; Grazioli, Fabiana; Esposito, Maria Luisa; Siani, Loredana; Traboni, Cinzia; Colloca, Stefano; Nicosia, Alfredo; Folgori, Antonella] ReiThera Srl Ex Okairos, I-00144 Rome, Italy; [Antrobus, Richard D.; Hill, Adrian; Barnes, Eleanor] Univ Oxford, Jenner Inst, Oxford OX3 7DQ, England; [Newell, Evan W.; Davis, Mark] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Newell, Evan W.] Singapore Immunol Network, Singapore 138648, Singapore; [Halliday, John; Kelly, Christabel; Klenerman, Paul; Barnes, Eleanor] Natl Inst Hlth Res, Oxford Biomed Res Ctr, Oxford OX3 7LE, England; [Halliday, John; Kelly, Christabel; Klenerman, Paul; Barnes, Eleanor] Translat Gastroenterol Unit, Oxford OX3 7LE, England; [Nicosia, Alfredo] CEINGE, I-80145 Naples, Italy; [Nicosia, Alfredo] Univ Naples Federico II, Dept Mol Med & Med Biotechnol, I-80131 Naples, Italy; [Cortese, Riccardo] Okairos AG, CH-4051 Basel, Switzerland[Swadling, Leo; Antrobus, Richard D.; Brown, Anthony; Richardson, Rachel; Halliday, John; Kelly, Christabel; Bowen, Dan; Fergusson, Joannah; Kurioka, Ayako; Hill, Adrian; Klenerman, Paul; Barnes, Eleanor] Univ Oxford, Nuffield Dept Med, Oxford OX1 3SY, England; [Capone, Stefania; Ammendola, Virginia; Del Sorbo, Mariarosaria; Grazioli, Fabiana; Esposito, Maria Luisa; Siani, Loredana; Traboni, Cinzia; Colloca, Stefano; Nicosia, Alfredo; Folgori, Antonella] ReiThera Srl Ex Okairos, I-00144 Rome, Italy; [Antrobus, Richard D.; Hill, Adrian; Barnes, Eleanor] Univ Oxford, Jenner Inst, Oxford OX3 7DQ, England; [Newell, Evan W.; Davis, Mark] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Newell, Evan W.] Singapore Immunol Network, Singapore 138648, Singapore; [Halliday, John; Kelly, Christabel; Klenerman, Paul; Barnes, Eleanor] Natl Inst Hlth Res, Oxford Biomed Res Ctr, Oxford OX3 7LE, England; [Halliday, John; Kelly, Christabel; Klenerman, Paul; Barnes, Eleanor] Translat Gastroenterol Unit, Oxford OX3 7LE, England; [Nicosia, Alfredo] CEINGE, I-80145 Naples, Italy; [Nicosia, Alfredo] Univ Naples Federico II, Dept Mol Med & Med Biotechnol, I-80131 Naples, Italy; [Cortese, Riccardo] Okairos AG, CH-4051 Basel, Switzerland	Barnes, E (reprint author), Univ Oxford, Nuffield Dept Med, Oxford OX1 3SY, England.	ellie.barnes@ndm.ox.ac.ukellie.barnes@ndm.ox.ac.uk	Newell, Evan/AAE-9470-2020; Newell, Evan W/F-9711-2012; Swadling, Leo/Y-5141-2019	Newell, Evan/0000-0002-2889-243X; Newell, Evan W/0000-0002-2889-243X; Swadling, Leo/0000-0002-0537-6715; Capone, Stefania/0000-0002-2272-120X; Barnes, Eleanor/0000-0002-0860-0831; Fergusson, Joannah/0000-0001-9223-1209	Medical Research Council (MRC) UKMedical Research Council UK (MRC); European Union (Framework VI; HEPACIVAC); MRC UK DCS (Developmental Clinical Studies) award; Oxford Martin Schools; National Institute for Health Research Oxford Biomedical Research CentreNational Institute for Health Research (NIHR); MRC CASE studentship; MRCMedical Research Council UK (MRC); Medical Research CouncilMedical Research Council UK (MRC) [MR/K010239/1]; National Institute for Health ResearchNational Institute for Health Research (NIHR) [NF-SI-0510-10204]	Supported by the Medical Research Council (MRC) UK and the European Union (Framework VI; HEPACIVAC) for funding the study and the manufacture of MVA-NSmut through an MRC UK DCS (Developmental Clinical Studies) award. E.B. is supported by the MRC as a Senior Clinical Fellow, the Oxford Martin Schools, and National Institute for Health Research Oxford Biomedical Research Centre. L. Swadling is supported by an MRC CASE studentship.	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Transl. Med.	NOV 5	2014	6	261							261ra153	10.1126/scitranslmed.3009185			16	Cell Biology; Medicine, Research & Experimental	Cell Biology; Research & Experimental Medicine	AS4GR	WOS:000344232400007	25378645	Green Accepted			2020-07-01	
J	Gaudilliere, B; Fragiadakis, GK; Bruggner, RV; Nicolau, M; Finck, R; Tingle, M; Silva, J; Ganio, EA; Yeh, CG; Maloney, WJ; Huddleston, JI; Goodman, SB; Davis, MM; Bendall, SC; Fantl, WJ; Angst, MS; Nolan, GP				Gaudilliere, Brice; Fragiadakis, Gabriela K.; Bruggner, Robert V.; Nicolau, Monica; Finck, Rachel; Tingle, Martha; Silva, Julian; Ganio, Edward A.; Yeh, Christine G.; Maloney, William J.; Huddleston, James I.; Goodman, Stuart B.; Davis, Mark M.; Bendall, Sean C.; Fantl, Wendy J.; Angst, Martin S.; Nolan, Garry P.			Clinical recovery from surgery correlates with single-cell immune signatures	SCIENCE TRANSLATIONAL MEDICINE			English	Article							NF-KAPPA-B; MASS CYTOMETRY; SEPTIC PATIENTS; EXPRESSION; STRESS; CONVALESCENCE; VALIDATION; PATHWAYS; OUTCOMES; SEPSIS	Delayed recovery from surgery causes personal suffering and substantial societal and economic costs. Whether immune mechanisms determine recovery after surgical trauma remains ill-defined. Single-cell mass cytometry was applied to serial whole-blood samples from 32 patients undergoing hip replacement to comprehensively characterize the phenotypic and functional immune response to surgical trauma. The simultaneous analysis of 14,000 phosphorylation events in precisely phenotyped immune cell subsets revealed uniform signaling responses among patients, demarcating a surgical immune signature. When regressed against clinical parameters of surgical recovery, including functional impairment and pain, strong correlations were found with STAT3 (signal transducer and activator of transcription), CREB (adenosine 3',5'-monophosphate response element-binding protein), and NF-kappa B (nuclear factor kappa B) signaling responses in subsets of CD14(+) monocytes (R = 0.7 to 0.8, false discovery rate <0.01). These sentinel results demonstrate the capacity of mass cytometry to survey the human immune system in a relevant clinical context. The mechanistically derived immune correlates point to diagnostic signatures, and potential therapeutic targets, that could postoperatively improve patient recovery.	[Gaudilliere, Brice; Tingle, Martha; Silva, Julian; Ganio, Edward A.; Yeh, Christine G.; Angst, Martin S.] Stanford Univ, Sch Med, Dept Anesthesiol Perioperat & Pain Med, Stanford, CA 94305 USA; [Gaudilliere, Brice; Fragiadakis, Gabriela K.; Bruggner, Robert V.; Nicolau, Monica; Finck, Rachel; Bendall, Sean C.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Fragiadakis, Gabriela K.; Finck, Rachel; Davis, Mark M.; Bendall, Sean C.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA; [Nicolau, Monica] Stanford Univ, Dept Math, Stanford, CA 94305 USA; [Nicolau, Monica] Stanford Univ, Ctr Canc Syst Biol, Stanford, CA 94305 USA; [Maloney, William J.; Huddleston, James I.; Goodman, Stuart B.] Stanford Univ, Dept Orthoped Surg, Redwood City, CA 94063 USA; [Fantl, Wendy J.] Stanford Univ, Dept Obstet & Gynecol, Stanford, CA 94305 USA[Gaudilliere, Brice; Tingle, Martha; Silva, Julian; Ganio, Edward A.; Yeh, Christine G.; Angst, Martin S.] Stanford Univ, Sch Med, Dept Anesthesiol Perioperat & Pain Med, Stanford, CA 94305 USA; [Gaudilliere, Brice; Fragiadakis, Gabriela K.; Bruggner, Robert V.; Nicolau, Monica; Finck, Rachel; Bendall, Sean C.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Fragiadakis, Gabriela K.; Finck, Rachel; Davis, Mark M.; Bendall, Sean C.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Bruggner, Robert V.] Stanford Univ, Biomed Informat Program, Stanford, CA 94305 USA; [Nicolau, Monica] Stanford Univ, Dept Math, Stanford, CA 94305 USA; [Nicolau, Monica] Stanford Univ, Ctr Canc Syst Biol, Stanford, CA 94305 USA; [Maloney, William J.; Huddleston, James I.; Goodman, Stuart B.] Stanford Univ, Dept Orthoped Surg, Redwood City, CA 94063 USA; [Fantl, Wendy J.] Stanford Univ, Dept Obstet & Gynecol, Stanford, CA 94305 USA	Angst, MS (reprint author), Stanford Univ, Sch Med, Dept Anesthesiol Perioperat & Pain Med, Stanford, CA 94305 USA.	ang@stanford.edu; gnolan@stanford.eduang@stanford.edu; gnolan@stanford.edu	Nolan, Garry/AAE-7903-2019		NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [T32GM089626, T32GM007276, 5T15LM007033-27, 1K99GM104148-01, UL1RR025744, 1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, 41000411217, 1U19AI100627, P01CA034233-22A1, U19AI057229, U54CA149145, S10RR027582-01]; Stanford Society of Physician Scholars grant; Stanford Bio-X graduate research fellowship; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; Food and Drug AdministrationUnited States Department of Health & Human Services [HHSF223201210194C]; National Cancer InstituteUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [U54 CA149145]; Stanford Department of Anesthesiology, Perioperative and Pain Medicine; California Institute for Regenerative MedicineCalifornia Institute for Regenerative Medicine [DR1-01477, RB2-01592]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; U.S. Department of DefenseUnited States Department of Defense [W81XWH-12-1-0591 OCRP-TIA NWC]	Supported by funds from NIH T32GM089626 and the Stanford Society of Physician Scholars grant (B.G.); the Stanford Bio-X graduate research fellowship and NIH T32GM007276 (G.K.F.); NIH 5T15LM007033-27 (R.V.B.); the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09) and NIH 1K99GM104148-01 (S.C.B.); the Food and Drug Administration Contract HHSF223201210194C and National Cancer Institute U54 CA149145 (M.N.); and the Stanford Department of Anesthesiology, Perioperative and Pain Medicine (M.S.A.). G.P.N. is supported by funds from NIH grants UL1RR025744, 1R01CA130826, 5U54CA143907, HHSN272200700038C, N01-HV-00242, 41000411217, 1U19AI100627, P01CA034233-22A1, U19AI057229, U54CA149145, and S10RR027582-01; the California Institute for Regenerative Medicine (DR1-01477 and RB2-01592); the European Commission (HEALTH.2010.1.2-1); and the U.S. Department of Defense (W81XWH-12-1-0591 OCRP-TIA NWC).	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Transl. Med.	SEP 24	2014	6	255							255ra131	10.1126/scitranslmed.3009701			12	Cell Biology; Medicine, Research & Experimental	Cell Biology; Research & Experimental Medicine	AR1BY	WOS:000343316800006	25253674	Green Accepted			2020-07-01	
J	Macaulay, IC; Ponting, CP; Voet, T				Macaulay, Iain C.; Ponting, Chris P.; Voet, Thierry			Single-Cell Multiomics: Multiple Measurements from Single Cells	TRENDS IN GENETICS			English	Review							GENOME-WIDE DETECTION; RNA-SEQ; MESSENGER-RNA; DNA-METHYLATION; MASS CYTOMETRY; TRANSCRIPTOME; PROTEINS; HETEROGENEITY; STEM; VARIABILITY	Single-cell sequencing provides information that is not confounded by genotypic or phenotypic heterogeneity of bulk samples. Sequencing of one molecular type (RNA, methylated DNA or open chromatin) in a single cell, furthermore, provides insights into the cell's phenotype and links to its genotype. Nevertheless, only by taking measurements of these phenotypes and genotypes from the same single cells can such inferences be made unambiguously. In this review, we survey the first experimental approaches that assay, in parallel, multiple molecular types from the same single cell, before considering the challenges and opportunities afforded by these and future technologies.	[Macaulay, Iain C.] Norwich Res Pk, Earlham Inst, Norwich NR4 7UH, Norfolk, England; [Ponting, Chris P.; Voet, Thierry] Wellcome Trust Sanger Inst, Sanger Inst EBI, Single Cell Genom Ctr, Hinxton CB10 1SA, England; [Ponting, Chris P.] Univ Edinburgh, MRC, MRC IGMM, Human Genet Unit, Crewe Rd, Edinburgh EH4 2XU, Midlothian, Scotland; [Voet, Thierry] Univ Leuven, Dept Human Genet, KU Leuven, B-3000 Leuven, Belgium[Macaulay, Iain C.] Norwich Res Pk, Earlham Inst, Norwich NR4 7UH, Norfolk, England; [Ponting, Chris P.; Voet, Thierry] Wellcome Trust Sanger Inst, Sanger Inst EBI, Single Cell Genom Ctr, Hinxton CB10 1SA, England; [Ponting, Chris P.] Univ Edinburgh, MRC, MRC IGMM, Human Genet Unit, Crewe Rd, Edinburgh EH4 2XU, Midlothian, Scotland; [Voet, Thierry] Univ Leuven, Dept Human Genet, KU Leuven, B-3000 Leuven, Belgium	Macaulay, IC (reprint author), Norwich Res Pk, Earlham Inst, Norwich NR4 7UH, Norfolk, England.; Ponting, CP; Voet, T (reprint author), Wellcome Trust Sanger Inst, Sanger Inst EBI, Single Cell Genom Ctr, Hinxton CB10 1SA, England.; Ponting, CP (reprint author), Univ Edinburgh, MRC, MRC IGMM, Human Genet Unit, Crewe Rd, Edinburgh EH4 2XU, Midlothian, Scotland.; Voet, T (reprint author), Univ Leuven, Dept Human Genet, KU Leuven, B-3000 Leuven, Belgium.	lain.Macaulay@earlham.ac.uk; Chris.Ponting@igmm.ed.ac.uk; thierry.voet@kuleuven.belain.Macaulay@earlham.ac.uk; Chris.Ponting@igmm.ed.ac.uk; thierry.voet@kuleuven.be	Voet, Thierry/E-8877-2017	Macaulay, Iain/0000-0002-6761-757X	Wellcome TrustWellcome Trust; BBSRCBiotechnology and Biological Sciences Research Council (BBSRC); MRCMedical Research Council UK (MRC); University of Leuven (KU Leuven, Belgium; SymBioSys) [PFV/10/016]; FWOFWO [G.0687.12, G.0924.15]; Medical Research CouncilMedical Research Council UK (MRC) [MC_PC_15075, MC_UU_12008/1]	Single-cell research and development at the Wellcome Trust Sanger Institute by C.P.P. and T.V. is funded by the Wellcome Trust. Single-cell technology development at the Earlham Institute by I.C.M. is funded by the BBSRC. C.P.P. research is also funded by the MRC. T.V. is also funded by the University of Leuven (KU Leuven, Belgium; SymBioSys, PFV/10/016) and the FWO (G.0687.12 and G.0924.15). We thank our colleagues at the WTSI/EBI Single Cell Genomics Centre for lively and helpful discussions.	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FEB	2017	33	2					155	168		10.1016/j.tig.2016.12.003			14	Genetics & Heredity	Genetics & Heredity	EK0RU	WOS:000393635600007	28089370	Green Published, Other Gold			2020-07-01	
J	Newell, EW; Sigal, N; Nair, N; Kidd, BA; Greenberg, HB; Davis, MM				Newell, Evan W.; Sigal, Natalia; Nair, Nitya; Kidd, Brian A.; Greenberg, Harry B.; Davis, Mark M.			Combinatorial tetramer staining and mass cytometry analysis facilitate T-cell epitope mapping and characterization	NATURE BIOTECHNOLOGY			English	Article							CLASS-I LIGANDS; MHC TETRAMERS; MEMORY; EXPRESSION; DIVERSITY; CONTINUUM; RESPONSES; PROTEIN; IMMUNE	It is currently not possible to predict which epitopes will be recognized by T cells in different individuals. This is a barrier to the thorough analysis and understanding of T-cell responses after vaccination or infection. Here, by combining mass cytometry with combinatorial peptide-MHC tetramer staining, we have developed a method allowing the rapid and simultaneous identification and characterization of T cells specific for many epitopes. We use this to screen up to 109 different peptide-MHC tetramers in a single human blood sample, while still retaining at least 23 labels to analyze other markers of T-cell phenotype and function. Among 77 candidate rotavirus epitopes, we identified six T-cell epitopes restricted to human leukocyte antigen (HLA)-A*0201 in the blood of healthy individuals. T cells specific for epitopes in the rotavirus VP3 protein displayed a distinct phenotype and were present at high frequencies in intestinal epithelium. This approach should be useful for the comprehensive analysis of T-cell responses to infectious diseases or vaccines.	[Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Newell, Evan W.; Nair, Nitya; Greenberg, Harry B.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Sigal, Natalia; Kidd, Brian A.; Greenberg, Harry B.; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Stanford, CA 94305 USA; [Nair, Nitya; Greenberg, Harry B.] Stanford Univ, Dept Med, Stanford, CA 94305 USA; [Davis, Mark M.] Stanford Univ, Howard Hughes Med Inst, Stanford, CA 94305 USA[Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Newell, Evan W.; Nair, Nitya; Greenberg, Harry B.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Sigal, Natalia; Kidd, Brian A.; Greenberg, Harry B.; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Stanford, CA 94305 USA; [Nair, Nitya; Greenberg, Harry B.] Stanford Univ, Dept Med, Stanford, CA 94305 USA; [Davis, Mark M.] Stanford Univ, Howard Hughes Med Inst, Stanford, CA 94305 USA	Davis, MM (reprint author), Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA.	evan_newell@immunol.a-star.edu.sg; mmdavis@stanford.eduevan_newell@immunol.a-star.edu.sg; mmdavis@stanford.edu	Nair, Nitya/D-2651-2017; Newell, Evan/AAE-9470-2020; Newell, Evan W/F-9711-2012	Nair, Nitya/0000-0002-9524-9313; Newell, Evan/0000-0002-2889-243X; Newell, Evan W/0000-0002-2889-243X; Kidd, Brian/0000-0003-2110-1145; greenberg, harry/0000-0002-2128-9080	Bill and Melinda Gates FoundationGates Foundation; National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19-AI057229, U19-AI090019]; Howard Hughes Medical InstituteHoward Hughes Medical Institute; American Cancer SocietyAmerican Cancer Society; Singapore Immunology NetworkAgency for Science Technology & Research (ASTAR)	We thank members of the Davis, Holden Maecker and Garry Nolan labs for sharing advice and experience concerning mass cytometry and antibody clone usage, especially M. Leipold for help with the mass cytometry instrument, and X. He, F. Wen, W. O'Gorman, A. Han, S. Bendall, O. Goldberger and Y.-H. Chien for helpful discussions. This work was supported by the Bill and Melinda Gates Foundation Grand Challenges Exploration phase I and II grants, National Institutes of Health grants U19-AI057229 and U19-AI090019, and The Howard Hughes Medical Institute. E.W.N. was supported by The American Cancer Society's Steven Stanley and Edward Albert Bielfelt Post-Doctoral Fellowship and by funding through the Singapore Immunology Network.	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Biotechnol.	JUL	2013	31	7					623	U81		10.1038/nbt.2593			9	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	180FS	WOS:000321579700019	23748502	Green Accepted			2020-07-01	
J	Brodin, P; Davis, MM				Brodin, Petter; Davis, Mark M.			Human immune system variation	NATURE REVIEWS IMMUNOLOGY			English	Review							DELTA T-CELLS; MASS CYTOMETRY; RHEUMATOID-ARTHRITIS; CIRCADIAN-RHYTHMS; PERIPHERAL-BLOOD; SEX-DIFFERENCES; EXPRESSION; DISEASE; AUTOIMMUNE; MICROBIOTA	The human immune system is highly variable between individuals but relatively stable over time within a given person. Recent conceptual and technological advances have enabled systems immunology analyses, which reveal the composition of immune cells and proteins in populations of healthy individuals. The range of variation and some specific influences that shape an individual's immune system is now becoming clearer. Human immune systems vary as a consequence of heritable and non-heritable influences, but symbiotic and pathogenic microbes and other non-heritable influences explain most of this variation. Understanding when and how such influences shape the human immune system is key for defining metrics of immunological health and understanding the risk of immune-mediated and infectious diseases.	[Brodin, Petter] Karolinska Inst, Dept Med Solna, Sci Life Lab, S-17165 Stockholm, Sweden; [Brodin, Petter] Karolinska Univ Hosp, Dept Neonatol, S-14186 Stockholm, Sweden; [Davis, Mark M.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94304 USA; [Davis, Mark M.] Stanford Univ, Sch Med, Inst Immun Transplantat & Infect, Stanford, CA 94304 USA; [Davis, Mark M.] Stanford Univ, Sch Med, Howard Hughes Med Inst, Stanford, CA 94304 USA[Brodin, Petter] Karolinska Inst, Dept Med Solna, Sci Life Lab, S-17165 Stockholm, Sweden; [Brodin, Petter] Karolinska Univ Hosp, Dept Neonatol, S-14186 Stockholm, Sweden; [Davis, Mark M.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94304 USA; [Davis, Mark M.] Stanford Univ, Sch Med, Inst Immun Transplantat & Infect, Stanford, CA 94304 USA; [Davis, Mark M.] Stanford Univ, Sch Med, Howard Hughes Med Inst, Stanford, CA 94304 USA	Brodin, P (reprint author), Karolinska Inst, Dept Med Solna, Sci Life Lab, S-17165 Stockholm, Sweden.; Brodin, P (reprint author), Karolinska Univ Hosp, Dept Neonatol, S-14186 Stockholm, Sweden.	petter.brodin@ki.sepetter.brodin@ki.se	Brodin, Petter/C-3317-2011	Brodin, Petter/0000-0002-8103-0046	European Research CouncilEuropean Research Council (ERC); Swedish Research CouncilSwedish Research Council; Swedish Society for Medical Research; Karolinska InstitutetKarolinska Institutet; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI090019, U19 AI057229]; Howard Hughes Medical InstituteHoward Hughes Medical Institute	P.B. is supported by a starting grant from the European Research Council, the Swedish Research Council, the Swedish Society for Medical Research, and Karolinska Institutet. M.M.D. is supported by NIH grants U19 AI090019, U19 AI057229 and the Howard Hughes Medical Institute.	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Whitacre CC, 2001, NAT IMMUNOL, V2, P777, DOI 10.1038/ni0901-777; Whitney AR, 2003, P NATL ACAD SCI USA, V100, P1896, DOI 10.1073/pnas.252784499; Wrammert J, 2008, NATURE, V453, P667, DOI 10.1038/nature06890; Xu GJ, 2015, SCIENCE, V348, DOI 10.1126/science.aaa0698; Ye CJ, 2014, SCIENCE, V345, P1311, DOI 10.1126/science.1254665	107	139	144	21	129	NATURE PUBLISHING GROUP	LONDON	MACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLAND	1474-1733	1474-1741		NAT REV IMMUNOL	Nat. Rev. Immunol.	JAN	2017	17	1					21	29		10.1038/nri.2016.125			9	Immunology	Immunology	EH6TX	WOS:000391907900009	27916977	Green Accepted			2020-07-01	
J	Dodd, D; Spitzer, MH; Van Treuren, W; Merrill, BD; Hryckowian, AJ; Higginbottom, SK; Le, A; Cowan, TM; Nolan, GP; Fischbach, MA; Sonnenburg, JL				Dodd, Dylan; Spitzer, Matthew H.; Van Treuren, William; Merrill, Bryan D.; Hryckowian, Andrew J.; Higginbottom, Steven K.; Le, Anthony; Cowan, Tina M.; Nolan, Garry P.; Fischbach, Michael A.; Sonnenburg, Justin L.			A gut bacterial pathway metabolizes aromatic amino acids into nine circulating metabolites	NATURE			English	Article							CLOSTRIDIUM-SPOROGENES; MASS CYTOMETRY; PRODUCTS; IMMUNE; BLOOD	The human gut microbiota produces dozens of metabolites that accumulate in the bloodstream(1,2), where they can have systemic effects on the host. Although these small molecules commonly reach concentrations similar to those achieved by pharmaceutical agents, remarkably little is known about the microbial metabolic pathways that produce them. Here we use a combination of genetics and metabolic profiling to characterize a pathway from the gut symbiont Clostridium sporogenes that generates aromatic amino acid metabolites. Our results reveal that this pathway produces twelve compounds, nine of which are known to accumulate in host serum. All three aromatic amino acids (tryptophan, phenylalanine and tyrosine) serve as substrates for the pathway, and it involves branching and alternative reductases for specific intermediates. By genetically manipulating C. sporogenes, we modulate serum levels of these metabolites in gnotobiotic mice, and show that in turn this affects intestinal permeability and systemic immunity. This work has the potential to provide the basis of a systematic effort to engineer the molecular output of the gut bacterial community.	[Dodd, Dylan; Spitzer, Matthew H.; Le, Anthony; Cowan, Tina M.] Stanford Univ, Sch Med, Dept Pathol, Stanford, CA 94305 USA; [Dodd, Dylan; Spitzer, Matthew H.; Van Treuren, William; Merrill, Bryan D.; Hryckowian, Andrew J.; Higginbottom, Steven K.; Nolan, Garry P.; Sonnenburg, Justin L.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Fischbach, Michael A.] Univ Calif San Francisco, California Inst Quantitat Biosci, San Francisco, CA 94143 USA; [Fischbach, Michael A.] Univ Calif San Francisco, Dept Bioengn & Therapeut Sci, San Francisco, CA 94143 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Parker Inst Canc Immunotherapy, Helen Diller Family Comprehens Canc Ctr, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Fischbach, Michael A.] Stanford Univ, Dept Bioengn & ChEM H, Stanford, CA 94305 USA[Dodd, Dylan; Spitzer, Matthew H.; Le, Anthony; Cowan, Tina M.] Stanford Univ, Sch Med, Dept Pathol, Stanford, CA 94305 USA; [Dodd, Dylan; Spitzer, Matthew H.; Van Treuren, William; Merrill, Bryan D.; Hryckowian, Andrew J.; Higginbottom, Steven K.; Nolan, Garry P.; Sonnenburg, Justin L.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Fischbach, Michael A.] Univ Calif San Francisco, California Inst Quantitat Biosci, San Francisco, CA 94143 USA; [Fischbach, Michael A.] Univ Calif San Francisco, Dept Bioengn & Therapeut Sci, San Francisco, CA 94143 USA; [Spitzer, Matthew H.] Univ Calif San Francisco, Parker Inst Canc Immunotherapy, Helen Diller Family Comprehens Canc Ctr, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Fischbach, Michael A.] Stanford Univ, Dept Bioengn & ChEM H, Stanford, CA 94305 USA	Sonnenburg, JL (reprint author), Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA.; Fischbach, MA (reprint author), Univ Calif San Francisco, California Inst Quantitat Biosci, San Francisco, CA 94143 USA.; Fischbach, MA (reprint author), Stanford Univ, Dept Bioengn & ChEM H, Stanford, CA 94305 USA.	fischbach@fischbachgroup.org; jsonnenburg@stanford.edufischbach@fischbachgroup.org; jsonnenburg@stanford.edu	Nolan, Garry/AAE-7903-2019; Merrill, Bryan/L-1235-2015	Merrill, Bryan/0000-0002-9967-5539; Fischbach, Michael/0000-0003-3079-8247; Spitzer, Matthew/0000-0002-5291-3819	National Institutes of Health NIDDKUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Diabetes & Digestive & Kidney Diseases (NIDDK) [R01-DK101674]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DP2-OD006515, U19AI057229, U19AI100627, R33CA183654, R33CA0183692, R01GM10983601, R01CA184968, R01CA19665701, R21CA183660, R01NS08953301, 5UH2AR067676, R01HL120724, DP5-OD023056]; FDAUnited States Department of Health & Human Services [BAA-12-00118]; Department of Defence [OC110674, W81XWH-14-1-0 180]; Gates FoundationGates Foundation [OPP1113682, DP1 DK113598, R01 DK110174]; HHMI-Simons Faculty Scholars Award; Byers Award in Basic Science; David and Lucile Packard FoundationThe David & Lucile Packard Foundation; BASF research grant; Burroughs Wellcome Investigators in the Pathogenesis of Infectious Disease Awards; NIH postdoctoral NRSA [T32-AI007328]; National Science FoundationNational Science Foundation (NSF) [DGE-114747]	We thank A. I. Scott for technical assistance and S. Yoshida (Kyoto University) for critical review of the manuscript. This work was funded by a grant from the National Institutes of Health NIDDK (R01-DK101674) to J.L.S. and M.A.F., an NIH Director's New Innovator Award (DP2-OD006515) to J.L.S., and Early Independence Award (DP5-OD023056) to M.H.S., FDA Grant BAA-12-00118 to G.P.N., NIH Awards U19AI057229, U19AI100627, R33CA183654, R33CA0183692, R01GM10983601, R01CA184968, R01CA19665701, R21CA183660, R01NS08953301, 5UH2AR067676 and R01HL120724 to G.P.N., Department of Defence Grants OC110674 and W81XWH-14-1-0 180 to G.P.N., Gates Foundation Grant OPP1113682 to G.P.N., DP1 DK113598 to M.A.F., R01 DK110174 to M.A.F., HHMI-Simons Faculty Scholars Award to M.A.F., Byers Award in Basic Science to M.A.F., a Fellowship for Science and Engineering from the David and Lucile Packard Foundation to M.A.F. and a BASF research grant to M.A.F, and two Burroughs Wellcome Investigators in the Pathogenesis of Infectious Disease Awards to M.A.F. and J.L.S. A.J.H. was supported by an NIH postdoctoral NRSA (T32-AI007328). W.V.T. and B.D.M. were each supported by a National Science Foundation Graduate Research Fellowship Grant No. DGE-114747.	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J	Mingueneau, M; Kreslavsky, T; Gray, D; Heng, T; Cruse, R; Ericson, J; Bendall, S; Spitzer, M; Nolan, G; Kobayashi, K; von Boehmer, H; Mathis, D; Benoist, C; Best, AJ; Knell, J; Goldrath, A; Jojic, V; Koller, D; Shay, T; Regev, A; Cohen, N; Brennan, P; Brenner, M; Kim, F; Rao, TN; Wagers, A; Heng, T; Ericson, J; Rothamel, K; Ortiz-Lopez, A; Mathis, D; Bezman, NA; Sun, JC; Min-Oo, G; Kim, CC; Lanier, LL; Miller, J; Brown, B; Merad, M; Gautier, EL; Jakubzick, C; Randolph, GJ; Monach, P; Blair, DA; Dustin, ML; Shinton, SA; Hardy, RR; Laidlaw, D; Collins, J; Gazit, R; Rossi, DJ; Malhotra, N; Sylvia, K; Kang, J; Kreslavsky, T; Fletcher, A; Elpek, K; Bellemare-Pelletier, A; Malhotra, D; Turley, S				Mingueneau, Michael; Kreslavsky, Taras; Gray, Daniel; Heng, Tracy; Cruse, Richard; Ericson, Jeffrey; Bendall, Sean; Spitzer, Matt; Nolan, Garry; Kobayashi, Koichi; von Boehmer, Harald; Mathis, Diane; Benoist, Christophe; Best, Adam J.; Knell, Jamie; Goldrath, Ananda; Jojic, Vladimir; Koller, Daphne; Shay, Tal; Regev, Aviv; Cohen, Nadia; Brennan, Patrick; Brenner, Michael; Kim, Francis; Rao, Tata Nageswara; Wagers, Amy; Heng, Tracy; Ericson, Jeffrey; Rothamel, Katherine; Ortiz-Lopez, Adriana; Mathis, Diane; Bezman, Natalie A.; Sun, Joseph C.; Min-Oo, Gundula; Kim, Charlie C.; Lanier, Lewis L.; Miller, Jennifer; Brown, Brian; Merad, Miriam; Gautier, Emmanuel L.; Jakubzick, Claudia; Randolph, Gwendalyn J.; Monach, Paul; Blair, David A.; Dustin, Michael L.; Shinton, Susan A.; Hardy, Richard R.; Laidlaw, David; Collins, Jim; Gazit, Roi; Rossi, Derrick J.; Malhotra, Nidhi; Sylvia, Katelyn; Kang, Joonsoo; Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemare-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon		Immunological Genome Consortium	The transcriptional landscape of alpha beta T cell differentiation	NATURE IMMUNOLOGY			English	Article							NEGATIVE SELECTION; POSITIVE SELECTION; GENE-EXPRESSION; PRE-TCR; C-MYC; CLONAL DELETION; LYMPHOCYTE DEVELOPMENT; INDUCED PROLIFERATION; THYMOCYTE SELECTION; CENTRAL TOLERANCE	The differentiation of abT cells from thymic precursors is a complex process essential for adaptive immunity. Here we exploited the breadth of expression data sets from the Immunological Genome Project to analyze how the differentiation of thymic precursors gives rise to mature T cell transcriptomes. We found that early T cell commitment was driven by unexpectedly gradual changes. In contrast, transit through the CD4(+)CD8(+) stage involved a global shutdown of housekeeping genes that is rare among cells of the immune system and correlated tightly with expression of the transcription factor c-Myc. Selection driven by major histocompatibility complex (MHC) molecules promoted a large-scale transcriptional reactivation. We identified distinct signatures that marked cells destined for positive selection versus apoptotic deletion. Differences in the expression of unexpectedly few genes accompanied commitment to the CD4(+) or CD8(+) lineage, a similarity that carried through to peripheral T cells and their activation, demonstrated by mass cytometry phosphoproteomics. The transcripts newly identified as encoding candidate mediators of key transitions help define the 'known unknowns' of thymocyte differentiation.	[Mingueneau, Michael; Cruse, Richard; Ericson, Jeffrey; Mathis, Diane; Benoist, Christophe; Heng, Tracy; Ericson, Jeffrey; Rothamel, Katherine; Ortiz-Lopez, Adriana; Mathis, Diane] Harvard Univ, Sch Med, Dept Microbiol & Immunol, Div Immunol, Boston, MA 02115 USA; [Kreslavsky, Taras; Kobayashi, Koichi; von Boehmer, Harald; Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemare-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon] Dana Farber Canc Inst, Boston, MA USA; [Gray, Daniel] Univ Melbourne, Dept Med Biol, Parkville, Vic 3052, Australia; [Gray, Daniel] Royal Melbourne Hosp, Walter & Eliza Hall Inst Med Res, Mol Genet Canc Div, Parkville, Vic 3050, Australia; [Gray, Daniel] Royal Melbourne Hosp, Walter & Eliza Hall Inst Med Res, Div Immunol, Parkville, Vic 3050, Australia; [Heng, Tracy] Monash Univ, Dept Anat & Dev Biol, Clayton, Vic, Australia; [Bendall, Sean; Spitzer, Matt; Nolan, Garry] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Palo Alto, CA 94304 USA; [Kobayashi, Koichi] Texas A&M Hlth Sci Ctr, Dept Microbial & Mol Pathogenesis, College Stn, TX USA; [Best, Adam J.; Knell, Jamie; Goldrath, Ananda] Univ Calif San Diego, Div Biol Sci, La Jolla, CA 92093 USA; [Jojic, Vladimir; Koller, Daphne] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Shay, Tal; Regev, Aviv] MIT, Broad Inst, Cambridge, MA 02139 USA; [Shay, Tal; Regev, Aviv] MIT, Dept Biol, Cambridge, MA USA; [Cohen, Nadia; Brennan, Patrick; Brenner, Michael] Brigham & Womens Hosp, Div Rheumatol Allergy & Immunol, Boston, MA 02115 USA; [Kim, Francis; Rao, Tata Nageswara; Wagers, Amy] Joslin Diabet Ctr, Boston, MA 02215 USA; [Bezman, Natalie A.; Sun, Joseph C.; Min-Oo, Gundula; Kim, Charlie C.; Lanier, Lewis L.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Miller, Jennifer; Brown, Brian; Merad, Miriam; Gautier, Emmanuel L.; Jakubzick, Claudia; Randolph, Gwendalyn J.] Mt Sinai Hosp, Icahn Med Inst, New York, NY 10029 USA; [Gautier, Emmanuel L.; Randolph, Gwendalyn J.] Washington Univ, Dept Pathol & Immunol, St Louis, MO USA; [Monach, Paul] Boston Univ, Dept Med, Boston, MA 02215 USA; [Blair, David A.; Dustin, Michael L.] NYU, Sch Med, Skirball Inst Biomol Med, New York, NY USA; [Shinton, Susan A.; Hardy, Richard R.] Fox Chase Canc Ctr, Philadelphia, PA 19111 USA; [Laidlaw, David] Brown Univ, Dept Comp Sci, Providence, RI 02912 USA; [Collins, Jim] Boston Univ, Howard Hughes Med Inst, Dept Biomed Engn, Boston, MA 02215 USA; [Gazit, Roi; Rossi, Derrick J.] Childrens Hosp, Program Mol Med, Boston, MA 02115 USA; [Malhotra, Nidhi; Sylvia, Katelyn; Kang, Joonsoo] Univ Massachusetts, Sch Med, Dept Pathol, Worcester, MA 01605 USA; [Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemare-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon] Harvard Univ, Sch Med, Boston, MA USA[Mingueneau, Michael; Cruse, Richard; Ericson, Jeffrey; Mathis, Diane; Benoist, Christophe; Heng, Tracy; Ericson, Jeffrey; Rothamel, Katherine; Ortiz-Lopez, Adriana; Mathis, Diane] Harvard Univ, Sch Med, Dept Microbiol & Immunol, Div Immunol, Boston, MA 02115 USA; [Kreslavsky, Taras; Kobayashi, Koichi; von Boehmer, Harald; Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemare-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon] Dana Farber Canc Inst, Boston, MA USA; [Gray, Daniel] Univ Melbourne, Dept Med Biol, Parkville, Vic 3052, Australia; [Gray, Daniel] Royal Melbourne Hosp, Walter & Eliza Hall Inst Med Res, Mol Genet Canc Div, Parkville, Vic 3050, Australia; [Gray, Daniel] Royal Melbourne Hosp, Walter & Eliza Hall Inst Med Res, Div Immunol, Parkville, Vic 3050, Australia; [Heng, Tracy] Monash Univ, Dept Anat & Dev Biol, Clayton, Vic, Australia; [Bendall, Sean; Spitzer, Matt; Nolan, Garry] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Palo Alto, CA 94304 USA; [Kobayashi, Koichi] Texas A&M Hlth Sci Ctr, Dept Microbial & Mol Pathogenesis, College Stn, TX USA; [Best, Adam J.; Knell, Jamie; Goldrath, Ananda] Univ Calif San Diego, Div Biol Sci, La Jolla, CA 92093 USA; [Jojic, Vladimir; Koller, Daphne] Stanford Univ, Dept Comp Sci, Stanford, CA 94305 USA; [Shay, Tal; Regev, Aviv] MIT, Broad Inst, Cambridge, MA 02139 USA; [Shay, Tal; Regev, Aviv] MIT, Dept Biol, Cambridge, MA USA; [Cohen, Nadia; Brennan, Patrick; Brenner, Michael] Brigham & Womens Hosp, Div Rheumatol Allergy & Immunol, Boston, MA 02115 USA; [Kim, Francis; Rao, Tata Nageswara; Wagers, Amy] Joslin Diabet Ctr, Boston, MA 02215 USA; [Bezman, Natalie A.; Sun, Joseph C.; Min-Oo, Gundula; Kim, Charlie C.; Lanier, Lewis L.] Univ Calif San Francisco, Dept Microbiol & Immunol, San Francisco, CA 94143 USA; [Miller, Jennifer; Brown, Brian; Merad, Miriam; Gautier, Emmanuel L.; Jakubzick, Claudia; Randolph, Gwendalyn J.] Mt Sinai Hosp, Icahn Med Inst, New York, NY 10029 USA; [Gautier, Emmanuel L.; Randolph, Gwendalyn J.] Washington Univ, Dept Pathol & Immunol, St Louis, MO USA; [Monach, Paul] Boston Univ, Dept Med, Boston, MA 02215 USA; [Blair, David A.; Dustin, Michael L.] NYU, Sch Med, Skirball Inst Biomol Med, New York, NY USA; [Shinton, Susan A.; Hardy, Richard R.] Fox Chase Canc Ctr, Philadelphia, PA 19111 USA; [Laidlaw, David] Brown Univ, Dept Comp Sci, Providence, RI 02912 USA; [Collins, Jim] Boston Univ, Howard Hughes Med Inst, Dept Biomed Engn, Boston, MA 02215 USA; [Gazit, Roi; Rossi, Derrick J.] Childrens Hosp, Program Mol Med, Boston, MA 02115 USA; [Malhotra, Nidhi; Sylvia, Katelyn; Kang, Joonsoo] Univ Massachusetts, Sch Med, Dept Pathol, Worcester, MA 01605 USA; [Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemare-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon] Harvard Univ, Sch Med, Boston, MA USA	Benoist, C (reprint author), Harvard Univ, Sch Med, Dept Microbiol & Immunol, Div Immunol, Boston, MA 02115 USA.	cbdm@hms.harvard.edu; cbdm@hms.harvard.educbdm@hms.harvard.edu; cbdm@hms.harvard.edu	Heng, Tracy/B-9847-2015; Nolan, Garry/AAE-7903-2019; Lanier, Lewis L/E-2139-2014; Mingueneau, Michael/O-5313-2019; Gazit, Roi/J-3352-2017; Dustin, Michael/AAM-4611-2020; Gray, Daniel HD/A-3293-2013; Gautier, Emmanuel L./E-2259-2017; Laidlaw, David/M-4686-2019	Heng, Tracy/0000-0003-4147-8986; Lanier, Lewis L/0000-0003-1308-3952; Mingueneau, Michael/0000-0002-3873-7329; Gray, Daniel HD/0000-0002-8457-8242; Gautier, Emmanuel L./0000-0003-2976-7566; Monach, Paul/0000-0003-4937-0515; Spitzer, Matthew/0000-0002-5291-3819; Dustin, Michael/0000-0003-4983-6389; Kreslavsky, Taras/0000-0002-6672-1914; Fletcher, Anne/0000-0003-1534-1142; Kim, Charles/0000-0001-6474-8227; kang, joonsoo/0000-0001-8419-7995; Miller, Jennifer/0000-0002-3591-369X; Gazit, Roi/0000-0002-0548-2147; Tata, Nageswara Rao/0000-0002-9928-5944; Malhotra, Deepali/0000-0002-8215-7639	US National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [AI072073]; Human Frontier Science ProgramHuman Frontier Science Program [HFSP-LT000096]; National Health and Medical Research CouncilNational Health and Medical Research Council of Australia [637353]; Australian Research CouncilAustralian Research Council [LP110201169]	We thank K. Hattori, A. Ortiz-Lopez and N. Asinovski for technical assistance with mice and antibodies; J. Moore and A. Kressler for flow cytometry; and C. Laplace for assistance with figure preparation. Supported by the US National Institutes of Health (AI072073), the Human Frontier Science Program (HFSP-LT000096 to M. M.), the National Health and Medical Research Council (637353 to D.G.) and the Australian Research Council (LP110201169 to, T.H.).	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Immunol.	JUN	2013	14	6					619	+		10.1038/ni.2590			16	Immunology	Immunology	146RG	WOS:000319107600015	23644507	Green Accepted			2020-07-01	
J	Shekhar, K; Brodin, P; Davis, MM; Chakraborty, AK				Shekhar, Karthik; Brodin, Petter; Davis, Mark M.; Chakraborty, Arup K.			Automatic Classification of Cellular Expression by Nonlinear Stochastic Embedding (ACCENSE)	PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA			English	Article						immunophenotyping; machine learning; class discovery; CyTOF; FACS	FLOW-CYTOMETRY DATA; T-CELLS; MEMORY; EFFECTOR; HOMEOSTASIS; LYMPHOCYTES; IMMUNE; CD4(+); GUIDE	Mass cytometry enables an unprecedented number of parameters to be measured in individual cells at a high throughput, but the large dimensionality of the resulting data severely limits approaches relying on manual "gating." Clustering cells based on phenotypic similarity comes at a loss of single-cell resolution and often the number of subpopulations is unknown a priori. Here we describe ACCENSE, a tool that combines nonlinear dimensionality reduction with density-based partitioning, and displays multivariate cellular phenotypes on a 2D plot. We apply ACCENSE to 35-parameter mass cytometry data from CD8(+) T cells derived from specific pathogen-free and germ-free mice, and stratify cells into phenotypic subpopulations. Our results show significant heterogeneity within the known CD8(+) T-cell subpopulations, and of particular note is that we find a large novel subpopulation in both specific pathogen-free and germ-free mice that has not been described previously. This subpopulation possesses a phenotypic signature that is distinct from conventional naive and memory subpopulations when analyzed by ACCENSE, but is not distinguishable on a biaxial plot of standard markers. We are able to automatically identify cellular subpopulations based on all proteins analyzed, thus aiding the full utilization of powerful new single-cell technologies such as mass cytometry.	[Shekhar, Karthik] MIT, Dept Chem Engn, Cambridge, MA 02139 USA; [Shekhar, Karthik; Chakraborty, Arup K.] Ragon Inst MGH MIT & Harvard, Boston, MA 02129 USA; [Brodin, Petter; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Sch Med, Stanford, CA 94304 USA; [Brodin, Petter; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Sch Med, Stanford, CA 94304 USA; [Chakraborty, Arup K.] MIT, Dept Chem Engn, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Phys, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Chem, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Biol Engn, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Inst Med Engn & Sci, Cambridge, MA 02139 USA[Shekhar, Karthik] MIT, Dept Chem Engn, Cambridge, MA 02139 USA; [Shekhar, Karthik; Chakraborty, Arup K.] Ragon Inst MGH MIT & Harvard, Boston, MA 02129 USA; [Brodin, Petter; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Sch Med, Stanford, CA 94304 USA; [Brodin, Petter; Davis, Mark M.] Stanford Univ, Inst Immun Transplantat & Infect, Sch Med, Stanford, CA 94304 USA; [Chakraborty, Arup K.] MIT, Dept Chem Engn, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Phys, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Chem, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Dept Biol Engn, Cambridge, MA 02139 USA; [Chakraborty, Arup K.] MIT, Inst Med Engn & Sci, Cambridge, MA 02139 USA	Davis, MM (reprint author), Stanford Univ, Dept Microbiol & Immunol, Sch Med, Stanford, CA 94304 USA.	mmdavis@stanford.edu; arupc@mit.edummdavis@stanford.edu; arupc@mit.edu	Brodin, Petter/C-3317-2011	Brodin, Petter/0000-0002-8103-0046	Poitras pre-doctoral fellowship; Ragon Institute of MGH; MIT; Harvard; Wenner-Gren Foundation; Swedish American Foundation [U189 AI 090019]; National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [PO1 AI091580]	We are grateful to Dr. G. Berman and Prof. J. Shaevitz for sharing unpublished work, and to Prof. A. Ferguson for a critical reading of the manuscript. We thank Prof. J. Sonnenburg for providing the GF mice. This research was supported by a Poitras pre-doctoral fellowship (K.S.), the Ragon Institute of MGH, MIT and Harvard (K.S. and A.K.C.), the Wenner-Gren Foundation and the Swedish American Foundation (P. B.), U189 AI 090019 (to M.M.D.), and a National Institutes of Health PO1 AI091580 (to A.K.C.).	Aghaeepour N, 2013, NAT METHODS, V10, P228, DOI [10.1038/NMETH.2365, 10.1038/nmeth.2365]; Amir ED, 2013, NAT BIOTECHNOL, V31, P545, DOI 10.1038/nbt.2594; Bendall SC, 2012, TRENDS IMMUNOL, V33, P323, DOI 10.1016/j.it.2012.02.010; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; CANTOR H, 1975, CELL IMMUNOL, V15, P180, DOI 10.1016/0008-8749(75)90174-4; Chung HC, 2012, CELL, V149, P1578, DOI 10.1016/j.cell.2012.04.037; Cover T. 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Natl. Acad. Sci. U. S. A.	JAN 7	2014	111	1					202	207		10.1073/pnas.1321405111			6	Multidisciplinary Sciences	Science & Technology - Other Topics	284VY	WOS:000329350700063	24344260	Green Published, Bronze			2020-07-01	
J	Frei, AP; Bava, FA; Zunder, ER; Hsieh, EWY; Chen, SY; Nolan, GP; Gherardini, PF				Frei, Andreas P.; Bava, Felice-Alessio; Zunder, Eli R.; Hsieh, Elena W. Y.; Chen, Shih-Yu; Nolan, Garry P.; Gherardini, Pier Federico			Highly multiplexed simultaneous detection of RNAs and proteins in single cells	NATURE METHODS			English	Article							IN-SITU HYBRIDIZATION; FLOW-CYTOMETRY; MESSENGER-RNA; GENE-EXPRESSION; INDIVIDUAL CELLS; PROXIMITY LIGATION; SEQ; HETEROGENEITY; COMPLEXES; LEVEL	To enable the detection of expression signatures specific to individual cells, we developed PLAYR (proximity ligation assay for RNA), a method for highly multiplexed transcript quantification by flow and mass cytometry that is compatible with standard antibody staining. When used with mass cytometry, PLAYR allowed for the simultaneous quantification of more than 40 different mRNAs and proteins. In primary cells, we quantified multiple transcripts, with the identity and functional state of each analyzed cell defined on the basis of the expression of a separate set of transcripts or proteins. By expanding high-throughput deep phenotyping of cells beyond protein epitopes to include RNA expression, PLAYR opens a new avenue for the characterization of cellular metabolism.	[Frei, Andreas P.; Bava, Felice-Alessio; Zunder, Eli R.; Hsieh, Elena W. Y.; Chen, Shih-Yu; Nolan, Garry P.; Gherardini, Pier Federico] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Zunder, Eli R.] Univ Virginia, Dept Biomed Engn, Charlottesville, VA USA; [Hsieh, Elena W. Y.] Univ Colorado, Dept Pediat, Denver, CO 80202 USA; [Hsieh, Elena W. Y.] Univ Colorado, Dept Immunol & Microbiol, Denver, CO 80202 USA[Frei, Andreas P.; Bava, Felice-Alessio; Zunder, Eli R.; Hsieh, Elena W. Y.; Chen, Shih-Yu; Nolan, Garry P.; Gherardini, Pier Federico] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Zunder, Eli R.] Univ Virginia, Dept Biomed Engn, Charlottesville, VA USA; [Hsieh, Elena W. Y.] Univ Colorado, Dept Pediat, Denver, CO 80202 USA; [Hsieh, Elena W. Y.] Univ Colorado, Dept Immunol & Microbiol, Denver, CO 80202 USA	Nolan, GP; Gherardini, PF (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	gnolan@stanford.edu; pfg@stanford.edugnolan@stanford.edu; pfg@stanford.edu	Chen, Shih-Yu/S-2919-2019; Nolan, Garry/AAE-7903-2019		Swiss National Science FoundationSwiss National Science Foundation (SNSF); EMBO Long-Term FellowshipEuropean Molecular Biology Organization (EMBO); Marie Curie International Outgoing FellowshipEuropean Union (EU); Human Frontier Science Program Long-Term FellowshipHuman Frontier Science Program; US National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, 1U19AI100627, R01CA184968, 1R33CA183654-01, R33CA183692, 1R01GM10983601, 1R21CA183660, 1R01NS08953301, OPP1113682, 5UH2AR067676, 1R01CA19665701, R01HL120724]; US Department of Defense Congressionally Directed Medical Research Programs [OC110674, 11491122]; Northrop-Grumman Corporation; Rachford & Carlotta A. Harris Endowed Chair	We thank L. Lanier (UCSF, San Francisco, California, USA) for providing the NKL cell line, and A. Trejo and A. Jager for technical assistance. A.P.F. is supported by a Fellowship for Prospective Researchers from the Swiss National Science Foundation, an EMBO Long-Term Fellowship and a Marie Curie International Outgoing Fellowship. P.F.G. is a Howard Hughes Medical Institute Fellow of the Life Sciences Research Foundation. F.-A.B. is supported by a Human Frontier Science Program Long-Term Fellowship. This work was supported by the US National Institutes of Health (grants U19 AI057229, 1U19AI100627, R01CA184968, 1R33CA183654-01, R33CA183692, 1R01GM10983601, 1R21CA183660, 1R01NS08953301, OPP1113682, 5UH2AR067676, 1R01CA19665701 and R01HL120724 to G.P.N.), US Department of Defense Congressionally Directed Medical Research Programs (grants OC110674 and 11491122 to G.P.N.), the Northrop-Grumman Corporation (G.P.N.) and the Rachford & Carlotta A. Harris Endowed Chair (G.P.N.).	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Methods	MAR	2016	13	3					269	+		10.1038/NMETH.3742			9	Biochemical Research Methods	Biochemistry & Molecular Biology	DF0NU	WOS:000371036700029	26808670	Green Accepted			2020-07-01	
J	Mrdjen, D; Pavlovic, A; Hartmann, FJ; Schreiner, B; Utz, SG; Leung, BP; Lelios, I; Heppner, FL; Kipnis, J; Merkler, D; Greter, M; Becher, B				Mrdjen, Dunja; Pavlovic, Anto; Hartmann, Felix J.; Schreiner, Bettina; Utz, Sebastian G.; Leung, Brian P.; Lelios, Iva; Heppner, Frank L.; Kipnis, Jonathan; Merkler, Doron; Greter, Melanie; Becher, Burkhard			High-Dimensional Single-Cell Mapping of Central Nervous System Immune Cells Reveals Distinct Myeloid Subsets in Health, Aging, and Disease	IMMUNITY			English	Article							TISSUE-RESIDENT MACROPHAGES; ALZHEIMERS-DISEASE; DENDRITIC CELLS; MASS CYTOMETRY; CHOROID-PLEXUS; GM-CSF; MULTIPLE-SCLEROSIS; STEM-CELLS; T-CELLS; MICROGLIA	Individual reports suggest that the central nervous system (CNS) contains multiple immune cell types with diverse roles in tissue homeostasis, immune defense, and neurological diseases. It has been challenging to map leukocytes across the entire brain, and in particular in pathology, where phenotypic changes and influx of blood-derived cells prevent a clear distinction between reactive leukocyte populations. Here, we applied high-dimensional single-cell mass and fluorescence cytometry, in parallel with genetic fate mapping systems, to identify, locate, and characterize multiple distinct immune populations within the mammalian CNS. Using this approach, we revealed that microglia, several subsets of border-associated macrophages and dendritic cells coexist in the CNS at steady state and exhibit disease-specific transformations in the immune microenvironment during aging and in models of Alzheimer's disease and multiple sclerosis. Together, these data and the described framework provide a resource for the study of disease mechanisms, potential biomarkers, and therapeutic targets in CNS disease.	[Mrdjen, Dunja; Pavlovic, Anto; Hartmann, Felix J.; Schreiner, Bettina; Utz, Sebastian G.; Leung, Brian P.; Lelios, Iva; Greter, Melanie; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, Zurich, Switzerland; [Heppner, Frank L.] Charite Univ Med Berlin, Dept Neuropathol, Berlin, Germany; [Kipnis, Jonathan] Univ Virginia, Ctr Brain Immunol & Glia, Dept Neurosci, Charlottesville, VA USA; [Merkler, Doron] Univ Geneva, Dept Pathol & Immunol, Geneva, Switzerland; [Merkler, Doron] Geneva Univ Hosp, Div Clin Pathol, Geneva, Switzerland[Mrdjen, Dunja; Pavlovic, Anto; Hartmann, Felix J.; Schreiner, Bettina; Utz, Sebastian G.; Leung, Brian P.; Lelios, Iva; Greter, Melanie; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, Zurich, Switzerland; [Heppner, Frank L.] Charite Univ Med Berlin, Dept Neuropathol, Berlin, Germany; [Kipnis, Jonathan] Univ Virginia, Ctr Brain Immunol & Glia, Dept Neurosci, Charlottesville, VA USA; [Merkler, Doron] Univ Geneva, Dept Pathol & Immunol, Geneva, Switzerland; [Merkler, Doron] Geneva Univ Hosp, Div Clin Pathol, Geneva, Switzerland	Becher, B (reprint author), Univ Zurich, Inst Expt Immunol, Zurich, Switzerland.	becher@immunology.uzh.chbecher@immunology.uzh.ch		Heppner, Frank/0000-0001-9816-8917; Greter, Melanie/0000-0002-7220-5369	Swiss national science foundationSwiss National Science Foundation (SNSF) [PP00P3_144781, 310030_146130, 316030_150768, 310030_170320]; European Union: FP7 ITN_NeuroKine; European Union: FP7 Project ATECT; University Priority Project Translational Cancer Research; National Science Foundation Graduate Research Fellowship ProgramNational Science Foundation (NSF) [DGE1418060]; Forschungskredit - University of Zurich	We thank the Mass- and Flow Cytometry Facility (University of Zurich) and the Center for Microscopy and Image Analysis (University of Zurich) for technical assistance; Lucy Robinson of Insight Editing London for critical review and editing of the manuscript; Nathalie Krakoski for discussions about BAMs; Nikolaus Deigendesch for APP/PS1 mouse transfers; Nir Yogev and Andrea Cogurra for discussions about all things meningeal; Will Macnair for comparison of protein and RNA expression; and Sarah Mundt for critical review and editing of the manuscript. This work was supported by grants from the Swiss national science foundation (PP00P3_144781 to M.G., 310030_146130, 316030_150768, 310030_170320 to B.B.), from the European Union: FP7 ITN_NeuroKine (B.B.) and FP7 Project ATECT (B.B.), from the University Priority Project Translational Cancer Research (B.B.), from the National Science Foundation Graduate Research Fellowship Program (DGE1418060) (B.P.L.), and the Forschungskredit awarded by the University of Zurich to D. Mrdjen.	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S	Sallusto, F		Littman, DR; Yokoyama, WM		Sallusto, Federica			Heterogeneity of Human CD4+T Cells Against Microbes	ANNUAL REVIEW OF IMMUNOLOGY, VOL 34	Annual Review of Immunology		English	Review; Book Chapter						human T cells; CD4+Th cells; cytokines; chemokine receptors; Mycobacteria; Candida	CD4(+) T-CELLS; CHEMOKINE RECEPTOR EXPRESSION; TRANSCRIPTION FACTOR PU.1; FOLLICULAR HELPER-CELLS; TH17 CELLS; TGF-BETA; SELECTIVE EXPRESSION; T-HELPER-2 CELLS; DENDRITIC CELLS; MASS CYTOMETRY	CD4(+) Thelper (Th) cells play a central role in the adaptive immune response by providing help to B cells and cytotoxic T cells and by releasing different types of cytokines in tissues to mediate protection against a wide range of pathogenic microorganisms. These functions are performed by different types of Th cells endowed with distinct migratory capacities and effector functions. Here we discuss how studies of the human T cell response to microbes have advanced our understanding of Th cell functional heterogeneity, in particular with the discovery of a distinct Th1 subset involved in the response to Mycobacteria and the characterization of two types of Th17 cells specific for extracellular bacteria or fungi. We also review new approaches to dissect at the clonal level the human CD4(+) T cell response induced by pathogens or vaccines that have revealed an unexpected degree of intraclonal diversification and propose a progressive and selective model of CD4(+) Tcell differentiation.	[Sallusto, Federica] Univ Svizzera Italiana, Ctr Med Immunol, CH-6500 Bellinzona, Switzerland; [Sallusto, Federica] Univ Svizzera Italiana, Cellular Immunol Lab, Inst Res Biomed, CH-6500 Bellinzona, Switzerland[Sallusto, Federica] Univ Svizzera Italiana, Ctr Med Immunol, CH-6500 Bellinzona, Switzerland; [Sallusto, Federica] Univ Svizzera Italiana, Cellular Immunol Lab, Inst Res Biomed, CH-6500 Bellinzona, Switzerland	Sallusto, F (reprint author), Univ Svizzera Italiana, Ctr Med Immunol, CH-6500 Bellinzona, Switzerland.; Sallusto, F (reprint author), Univ Svizzera Italiana, Cellular Immunol Lab, Inst Res Biomed, CH-6500 Bellinzona, Switzerland.	federica.sallusto@irb.usi.chfederica.sallusto@irb.usi.ch					Acosta-Rodriguez EV, 2007, NAT IMMUNOL, V8, P639, DOI 10.1038/ni1467; Annunziato F, 2001, J ALLERGY CLIN IMMUN, V108, P815; Annunziato F, 2007, J EXP MED, V204, P1849, DOI 10.1084/jem.20070663; Anuradha R, 2013, J IMMUNOL, V191, P2466, DOI 10.4049/jimmunol.1300911; Arlehamn CL, 2014, J IMMUNOL, V193, P2931, DOI 10.4049/jimmunol.1401151; 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Rev. Immunol.		2016	34						317	334		10.1146/annurev-immunol-032414-112056			18	Immunology	Immunology	BE7XH	WOS:000376011300012	27168241				2020-07-01	
J	Lanni, EJ; Rubakhin, SS; Sweedler, JV				Lanni, Eric J.; Rubakhin, Stanislav S.; Sweedler, Jonathan V.			Mass spectrometry imaging and profiling of single cells	JOURNAL OF PROTEOMICS			English	Review						Secondary ion mass spectrometry; Matrix assisted laser desorption/ionization; Subcellular profiling; Elemental imaging; Mass cytometry; Tissue imaging	LASER-DESORPTION IONIZATION; PRIMARY ION-BOMBARDMENT; LA-ICP-MS; SECONDARY-ION; TOF-SIMS; MALDI-MS; CHEMICAL-COMPOSITION; SAMPLE PREPARATION; TUMOR-CELLS; RELATIVE QUANTIFICATION	Mass spectrometry imaging and profiling of individual cells and subcellular structures provide unique analytical capabilities for biological and biomedical research, including determination of the biochemical heterogeneity of cellular populations and intracellular localization of pharmaceuticals. Two mass spectrometry technologies-secondary ion mass spectrometry (SIMS) and matrix assisted laser desorption/ionization mass spectrometry (MALDI MS)-are most often used in micro-bioanalytical investigations. Recent advances in ion probe technologies have increased the dynamic range and sensitivity of analyte detection by SIMS, allowing two- and three-dimensional localization of analytes in a variety of cells. SIMS operating in the mass spectrometry imaging (MSI) mode can routinely reach spatial resolutions at the submicron level; therefore, it is frequently used in studies of the chemical composition of subcellular structures. MALDI MS offers a large mass range and high sensitivity of analyte detection. It has been successfully applied in a variety of single-cell and organelle profiling studies. Innovative instrumentation such as scanning microprobe MALDI and mass microscope spectrometers enables new subcellular MSI measurements. Other approaches for MS-based chemical imaging and profiling include those based on near-field laser ablation and inductively-coupled plasma MS analysis, which offer complementary capabilities for subcellular chemical imaging and profiling. This article is part of a Special Issue entitled: Imaging Mass Spectrometry: A User's Guide to a New Technique for Biological and Biomedical Research. (C) 2012 Elsevier B.V. All rights reserved.	[Sweedler, Jonathan V.] Univ Illinois, Dept Chem, Urbana, IL 61801 USA; Univ Illinois, Beckman Inst Sci & Technol, Urbana, IL 61801 USA[Sweedler, Jonathan V.] Univ Illinois, Dept Chem, Urbana, IL 61801 USA; Univ Illinois, Beckman Inst Sci & Technol, Urbana, IL 61801 USA	Sweedler, JV (reprint author), Univ Illinois, Dept Chem, 1209 W Calif St, Urbana, IL 61801 USA.	jsweedle@illinois.edujsweedle@illinois.edu	Rubakhin, Stanislav S/E-6685-2017; Sweedler, Jonathan V/A-9405-2009	Rubakhin, Stanislav S/0000-0003-0437-1493; Sweedler, Jonathan V/0000-0003-3107-9922	National Institute on Drug Abuse (NIDA)United States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute on Drug Abuse (NIDA) [P30 DA018310]; Department of EnergyUnited States Department of Energy (DOE) [DE-SC0006642]	This work was supported by Award No. P30 DA018310 from the National Institute on Drug Abuse (NIDA) and the Department of Energy by Award No. DE-SC0006642. The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies. We also thank Stephanie Baker for assistance with manuscript preparation and reviewers for helpful feedback. The authors declare they have no conflicts of interest.	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Proteomics	AUG 30	2012	75	16			SI		5036	5051		10.1016/j.jprot.2012.03.017			16	Biochemical Research Methods	Biochemistry & Molecular Biology	002ID	WOS:000308524400014	22498881	Green Accepted			2020-07-01	
J	Spitzer, MH; Gherardini, PF; Fragiadakis, GK; Bhattacharya, N; Yuan, RT; Hotson, AN; Finck, R; Carmi, Y; Zunder, ER; Fantl, WJ; Bendall, SC; Engleman, EG; Nolan, GP				Spitzer, Matthew H.; Gherardini, Pier Federico; Fragiadakis, Gabriela K.; Bhattacharya, Nupur; Yuan, Robert T.; Hotson, Andrew N.; Finck, Rachel; Carmi, Yaron; Zunder, Eli R.; Fantl, Wendy J.; Bendall, Sean C.; Engleman, Edgar G.; Nolan, Garry P.			An interactive reference framework for modeling a dynamic immune system	SCIENCE			English	Article							MASS CYTOMETRY; GENE-EXPRESSION; T-CELLS; MOUSE; NETWORK; IDENTIFICATION; PROGRESSION; VISUALIZATION; BIOLOGY; GAMMA	Immune cells function in an interacting hierarchy that coordinates the activities of various cell types according to genetic and environmental contexts. We developed graphical approaches to construct an extensible immune reference map from mass cytometry data of cells from different organs, incorporating landmark cell populations as flags on the map to compare cells from distinct samples. The maps recapitulated canonical cellular phenotypes and revealed reproducible, tissue-specific deviations. The approach revealed influences of genetic variation and circadian rhythms on immune system structure, enabled direct comparisons of murine and human blood cell phenotypes, and even enabled archival fluorescence-based flow cytometry data to be mapped onto the reference framework. This foundational reference map provides a working definition of systemic immune organization to which new data can be integrated to reveal deviations driven by genetics, environment, or pathology.	[Spitzer, Matthew H.; Gherardini, Pier Federico; Fragiadakis, Gabriela K.; Hotson, Andrew N.; Finck, Rachel; Zunder, Eli R.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Bhattacharya, Nupur; Yuan, Robert T.; Carmi, Yaron; Bendall, Sean C.; Engleman, Edgar G.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Yuan, Robert T.; Bendall, Sean C.; Engleman, Edgar G.; Nolan, Garry P.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Fantl, Wendy J.] Stanford Univ, Dept Obstet & Gynecol, Div Gynecol Oncol, Stanford, CA 94305 USA[Spitzer, Matthew H.; Gherardini, Pier Federico; Fragiadakis, Gabriela K.; Hotson, Andrew N.; Finck, Rachel; Zunder, Eli R.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Bhattacharya, Nupur; Yuan, Robert T.; Carmi, Yaron; Bendall, Sean C.; Engleman, Edgar G.] Stanford Univ, Dept Pathol, Stanford, CA 94305 USA; [Spitzer, Matthew H.; Yuan, Robert T.; Bendall, Sean C.; Engleman, Edgar G.; Nolan, Garry P.] Stanford Univ, Program Immunol, Stanford, CA 94305 USA; [Fantl, Wendy J.] Stanford Univ, Dept Obstet & Gynecol, Div Gynecol Oncol, Stanford, CA 94305 USA	Spitzer, MH (reprint author), Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA.	matthew.spitzer@stanford.edu; gnolan@stanford.edumatthew.spitzer@stanford.edu; gnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Spitzer, Matthew/0000-0002-5291-3819	Fluidigm Sciences; George D. Smith Stanford graduate fellowship; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [F31CA189331, T32GM007276, NRSA F32 GM093508-01, K99GM104148-01, 1U19AI100627, 1R01GM109836, 7500108142]; Stanford Bio-X graduate fellowship; CIRM Basic Biology II [RB2-01592]; Damon Runyon Cancer Research Foundation [DRG-2017-09]; NIAIDUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [HHSN272201200028C, PN2EY018228 0158 G KB065, 1R01CA130826]; DODUnited States Department of Defense [OC110674, 11491122]; Bill and Melinda Gates FoundationGates Foundation [OPP1113682]; CIRMCalifornia Institute for Regenerative Medicine [RB2-01592, DR1-01477]; FDAUnited States Department of Health & Human Services [HHSF223201210194C BAA-12-00118]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; Rachford and Carlota A. Harris endowed professorship; The NIAID [5U54CA143907NIH, HHSN272200700038C, N01-HV-00242, 41000411217, 5-24927, P01 CA034233-22A1, RFA CA 09-009, RFA CA 09-011, U19 AI057229, U54CA149145, 5R01AI073724, R01CA184968, R33 CA183654, R33 CA183692, 1R01NS089533, 201303028]	All mass cytometry data are accessible at www.cytobank.org/nolanlab/reports and the. R package is available at https://github.com/nolanlab. R. Finck, S. Bendall, and G.P.N. have served as paid consultants of Fluidigm Sciences, the maker of the mass cytometry instrumentation and reagents used for the data collection in this study. We thank J. Kenkel, B. Burt, D.-H. Wang, and M. Ch'ng for their assistance in tissue processing; A. Trejo and A. Jager for mass cytometry quality control and maintenance; B. Gaudilliere and M. Angst for access to human whole blood data; and M. Angelo, C. Loh, N. Reticker-Flynn, and L. Sanman for constructive feedback. Supported by a George D. Smith Stanford graduate fellowship and NIH grant F31CA189331 (M.H.S.); a Stanford Bio-X graduate fellowship and NIH grant T32GM007276 (G.K.F.); CIRM Basic Biology II RB2-01592 and NIH grant NRSA F32 GM093508-01 (E.R.Z.); Damon Runyon Cancer Research Foundation fellowship DRG-2017-09 and NIH grant K99GM104148-01 (S. C. B.); NIH grants 1U19AI100627, 1R01GM109836, and 7500108142, NIAID Bioinformatics Support Contract HHSN272201200028C, PN2EY018228 0158 G KB065, 1R01CA130826, 5U54CA143907NIH, HHSN272200700038C, N01-HV-00242, 41000411217, 5-24927, P01 CA034233-22A1, RFA CA 09-009, RFA CA 09-011, U19 AI057229, U54CA149145, 5R01AI073724, R01CA184968, R33 CA183654, R33 CA183692, 1R01NS089533, 201303028; DOD grants OC110674 and 11491122; Bill and Melinda Gates Foundation grant OPP1113682; CIRM grants DR1-01477 and RB2-01592; FDA grant HHSF223201210194C BAA-12-00118; European Commission grant HEALTH.2010.1.2-1; and the Rachford and Carlota A. Harris endowed professorship (G.P.N.).. P.F.G. is a Howard Hughes Medical Institute Fellow of the Life Sciences Research Foundation.	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J	Angerer, P; Haghverdi, L; Buttner, M; Theis, FJ; Marr, C; Buettner, F				Angerer, Philipp; Haghverdi, Laleh; Buettner, Maren; Theis, Fabian J.; Marr, Carsten; Buettner, Florian			destiny: diffusion maps for large-scale single cell data in R	BIOINFORMATICS			English	Article								Diffusion maps are a spectral method for non-linear dimension reduction and have recently been adapted for the visualization of single-cell expression data. Here we present destiny, an efficient R implementation of the diffusion map algorithm. Our package includes a single-cell specific noise model allowing for missing and censored values. In contrast to previous implementations, we further present an efficient nearest-neighbour approximation that allows for the processing of hundreds of thousands of cells and a functionality for projecting new data on existing diffusion maps. We exemplarily apply destiny to a recent time-resolved mass cytometry dataset of cellular reprogramming.	[Angerer, Philipp; Haghverdi, Laleh; Buettner, Maren; Theis, Fabian J.; Marr, Carsten; Buettner, Florian] Helmholtz Zentrum Munchen, German Res Ctr Environm Hlth, Inst Computat Biol, Ingolstadter Landstr 1, D-85764 Neuherberg, Germany; [Theis, Fabian J.] Tech Univ Munich, Ctr Math, Chair Math Modeling Biol Syst, Boltzmannstr 3, D-85748 Garching, Germany[Angerer, Philipp; Haghverdi, Laleh; Buettner, Maren; Theis, Fabian J.; Marr, Carsten; Buettner, Florian] Helmholtz Zentrum Munchen, German Res Ctr Environm Hlth, Inst Computat Biol, Ingolstadter Landstr 1, D-85764 Neuherberg, Germany; [Theis, Fabian J.] Tech Univ Munich, Ctr Math, Chair Math Modeling Biol Syst, Boltzmannstr 3, D-85748 Garching, Germany	Marr, C; Buettner, F (reprint author), Helmholtz Zentrum Munchen, German Res Ctr Environm Hlth, Inst Computat Biol, Ingolstadter Landstr 1, D-85764 Neuherberg, Germany.	carsten.marr@helmholtz-muenchen.de; f.buettner@helmholtz-muenchen.decarsten.marr@helmholtz-muenchen.de; f.buettner@helmholtz-muenchen.de	Marr, Carsten/B-3696-2013; Angerer, Philipp/L-5236-2019	Marr, Carsten/0000-0003-2154-4552; Angerer, Philipp/0000-0002-0369-2888; Buttner, Maren/0000-0002-6189-3792; Theis, Fabian/0000-0002-2419-1943; Haghverdi, Laleh/0000-0001-9280-9170; Buettner, Florian/0000-0001-5587-6761	UK Medical Research CouncilMedical Research Council UK (MRC); Bavarian Research Network for Molecular Biosystems (BioSysNet); ERCEuropean Research Council (ERC); DFG Fellowship through the Graduate School of Quantitative Biosciences Munich (QBM); Medical Research CouncilMedical Research Council UK (MRC) [MR/M01536X/1]	Supported by the UK Medical Research Council (Career Development Award to FB), the Bavarian Research Network for Molecular Biosystems (BioSysNet) and the ERC (starting grant LatentCauses to FJT). MP, is supported by a DFG Fellowship through the Graduate School of Quantitative Biosciences Munich (QBM).	Buettner F, 2012, BIOINFORMATICS, V28, pI626, DOI 10.1093/bioinformatics/bts385; Coifman RR, 2008, MULTISCALE MODEL SIM, V7, P842, DOI 10.1137/070696325; Coifman RR, 2005, P NATL ACAD SCI USA, V102, P7426, DOI 10.1073/pnas.0500334102; Haghverdi L, 2015, BIOINFORMATICS, V31, P2989, DOI 10.1093/bioinformatics/btv325; Richards J., 2014, DIFFUSIONMAP DIFFUSI; Roditi LD, 2015, CURR OPIN BIOTECH, V34, P9, DOI 10.1016/j.copbio.2014.10.010; Zunder ER, 2015, CELL STEM CELL, V16, P323, DOI 10.1016/j.stem.2015.01.015	7	114	113	4	11	OXFORD UNIV PRESS	OXFORD	GREAT CLARENDON ST, OXFORD OX2 6DP, ENGLAND	1367-4803	1460-2059		BIOINFORMATICS	Bioinformatics	APR 15	2016	32	8					1241	1243		10.1093/bioinformatics/btv715			3	Biochemical Research Methods; Biotechnology & Applied Microbiology; Computer Science, Interdisciplinary Applications; Mathematical & Computational Biology; Statistics & Probability	Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology; Computer Science; Mathematical & Computational Biology; Mathematics	DJ8QJ	WOS:000374476800023	26668002	Bronze, Green Published			2020-07-01	
J	Lin, JR; Fallahi-Sichani, M; Sorger, PK				Lin, Jia-Ren; Fallahi-Sichani, Mohammad; Sorger, Peter K.			Highly multiplexed imaging of single cells using a high-throughput cyclic immunofluorescence method	NATURE COMMUNICATIONS			English	Article							TYRAMIDE SIGNAL AMPLIFICATION; PRIMARY ANTIBODIES; IMMUNOHISTOCHEMISTRY; VISUALIZATION; HETEROGENEITY; POPULATIONS; RESPONSES; REVEALS	Single-cell analysis reveals aspects of cellular physiology not evident from population-based studies, particularly in the case of highly multiplexed methods such as mass cytometry (CyTOF) able to correlate the levels of multiple signalling, differentiation and cell fate markers. Immunofluorescence (IF) microscopy adds information on cell morphology and the microenvironment that are not obtained using flow-based techniques, but the multiplicity of conventional IF is limited. This has motivated development of imaging methods that require specialized instrumentation, exotic reagents or proprietary protocols that are difficult to reproduce in most laboratories. Here we report a public-domain method for achieving high multiplicity single-cell IF using cyclic immunofluorescence (CycIF), a simple and versatile procedure in which four-colour staining alternates with chemical inactivation of fluorophores to progressively build a multichannel image. Because CycIF uses standard reagents and instrumentation and is no more expensive than conventional IF, it is suitable for high-throughput assays and screening applications.	[Lin, Jia-Ren; Sorger, Peter K.] Harvard Univ, Sch Med, HMS LINCS Ctr, Boston, MA 02115 USA; [Lin, Jia-Ren; Sorger, Peter K.] Harvard Univ, Sch Med, Lab Syst Pharmacol, Boston, MA 02115 USA; [Fallahi-Sichani, Mohammad; Sorger, Peter K.] Harvard Univ, Sch Med, Dept Syst Biol, Boston, MA 02115 USA[Lin, Jia-Ren; Sorger, Peter K.] Harvard Univ, Sch Med, HMS LINCS Ctr, Boston, MA 02115 USA; [Lin, Jia-Ren; Sorger, Peter K.] Harvard Univ, Sch Med, Lab Syst Pharmacol, Boston, MA 02115 USA; [Fallahi-Sichani, Mohammad; Sorger, Peter K.] Harvard Univ, Sch Med, Dept Syst Biol, Boston, MA 02115 USA	Sorger, PK (reprint author), Harvard Univ, Sch Med, HMS LINCS Ctr, Boston, MA 02115 USA.	peter_sorger@hms.harvard.edupeter_sorger@hms.harvard.edu		Sorger, Peter/0000-0002-3364-1838	NIH LINCS grant [U54-HL127365]	We thank J. Albeck and S. Sampattavanich for the FOXO-GFP reporter, GE for early access to a CyTell imaging system and E. Williams for the CycIF web-page. This work was funded by NIH LINCS grant U54-HL127365. M.F. is a Merck Fellow of the Life Sciences Research Foundation.	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J., 2014, CYTOM PART A, V87, P558; STERNBERGER LA, 1986, J HISTOCHEM CYTOCHEM, V34, P599, DOI 10.1177/34.5.3517144; Thorne CA, 2015, NAT CHEM BIOL, V11, P58, DOI 10.1038/nchembio.1690; Tornehave D, 2000, HISTOCHEM CELL BIOL, V113, P19; Toth ZE, 2007, J HISTOCHEM CYTOCHEM, V55, P545, DOI 10.1369/jhc.6A7134.2007; Ullal AV, 2014, SCI TRANSL MED, V6, DOI 10.1126/scitranslmed.3007361; Zrazhevskiy P, 2013, NAT COMMUN, V4, DOI 10.1038/ncomms2635	31	114	115	3	41	NATURE PUBLISHING GROUP	LONDON	MACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLAND	2041-1723			NAT COMMUN	Nat. Commun.	SEP	2015	6								8390	10.1038/ncomms9390			7	Multidisciplinary Sciences	Science & Technology - Other Topics	CT9KY	WOS:000363137100005	26399630	DOAJ Gold, Green Published, Green Accepted			2020-07-01	
J	Behbehani, GK; Bendall, SC; Clutter, MR; Fantl, WJ; Nolan, GP				Behbehani, Gregory K.; Bendall, Sean C.; Clutter, Matthew R.; Fantl, Wendy J.; Nolan, Garry P.			Single-cell mass cytometry adapted to measurements of the cell cycle	CYTOMETRY PART A			English	Article						mass cytometry; cell cycle; flow cytometry; retinoblastoma; iododeoxyuridine; hematopoiesis	HEMATOPOIETIC STEM; HISTONE H3; PROLIFERATION; EXPRESSION; LEUKEMIA	Mass cytometry is a recently introduced technology that utilizes transition element isotope-tagged antibodies for protein detection on a single-cell basis. By circumventing the limitations of emission spectral overlap associated with fluorochromes utilized in traditional flow cytometry, mass cytometry currently allows measurement of up to 40 parameters per cell. Recently, a comprehensive mass cytometry analysis was described for the hematopoietic differentiation program in human bone marrow from a healthy donor. The current study describes approaches to delineate cell cycle stages utilizing 5-iodo-2-deoxyuridine (IdU) to mark cells in S phase, simultaneously with antibodies against cyclin B1, cyclin A, and phosphorylated histone H3 (S28) that characterize the other cell cycle phases. Protocols were developed in which an antibody against phosphorylated retinoblastoma protein (Rb) at serines 807 and 811 was used to separate cells in G0 and G1 phases of the cell cycle. This mass cytometry method yielded cell cycle distributions of both normal and cancer cell populations that were equivalent to those obtained by traditional fluorescence cytometry techniques. We applied this to map the cell cycle phases of cells spanning the hematopoietic hierarchy in healthy human bone marrow as a prelude to later studies with cancers and other disorders of this lineage. (C) 2012 International Society for Advancement of Cytometry	[Behbehani, Gregory K.; Bendall, Sean C.; Clutter, Matthew R.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Sch Med, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Hematol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Oncol, Stanford, CA 94305 USA[Behbehani, Gregory K.; Bendall, Sean C.; Clutter, Matthew R.; Fantl, Wendy J.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Sch Med, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Hematol, Stanford, CA 94305 USA; [Behbehani, Gregory K.] Stanford Univ, Sch Med, Div Oncol, Stanford, CA 94305 USA	Behbehani, GK (reprint author), 269 Campus Dr,CCSR 3205, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019		NIH/NCIUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, NCI RFA CA 09-011, NHLBI-HV-1005, CIRM DR1-01477, RB2-01592]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; Bill and Melinda Gates FoundationGates Foundation [GF12141-137101]; Rachford and Carlota A. Harris Endowed Professorship; Stanford Cancer Center	Grant sponsor: NIH/NCI; Grant number: U19 AI057229; Grant number: P01 CA034233; Grant number: HHSN272200700038C; Grant number: 1R01CA130826; Grant number: NCI RFA CA 09-011; Grant number: NHLBI-HV-1005(2); Grant number: CIRM DR1-01477; Grant number: RB2-01592; Grant sponsor: European Commission; Grant number: HEALTH. 2010.1.2-1; Grant sponsor: Bill and Melinda Gates Foundation; Grant number: GF12141-137101; Grant sponsor: Rachford and Carlota A. Harris Endowed Professorship (G.P.N.); Grant sponsor: Stanford Cancer Center (Developmental Research Grant, G.K.B.);	Bandura DR, 2009, ANAL CHEM, V81, P6813, DOI 10.1021/ac901049w; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Bryder D, 2006, AM J PATHOL, V169, P338, DOI 10.2353/ajpath.2006.060312; Chao MP, 2008, COLD SH Q B, V73, P439, DOI 10.1101/sqb.2008.73.031; Cheshier SH, 2007, STEM CELLS DEV, V16, P707, DOI 10.1089/scd.2007.0017; Cheshier SP, 1999, P NATL ACAD SCI USA, V96, P3120, DOI 10.1073/pnas.96.6.3120; Darzynkiewicz Z, 2004, CYTOM PART A, V58A, P21, DOI 10.1002/cyto.a.20003; Darzynkiewicz Z, 1996, CYTOMETRY, V25, P1; Darzynkiewicz Z., 2004, CURR PROTOC CYTOM, DOI DOI 10.1002/0471142956.CY0703S30; Dey-Guha I, 2011, P NATL ACAD SCI USA, V108, P12845, DOI 10.1073/pnas.1109632108; GONG JP, 1994, CANCER RES, V54, P4285; Goto H, 2002, GENES CELLS, V7, P11, DOI 10.1046/j.1356-9597.2001.00498.x; GRIFFIN JD, 1986, BLOOD, V68, P1185; Hahne F, 2009, BMC BIOINFORMATICS, V10, DOI 10.1186/1471-2105-10-106; Head DR, 2011, BONE MARROW RES, DOI 10.1155/2011/950934; Hirata A, 2004, J HISTOCHEM CYTOCHEM, V52, P1503, DOI 10.1369/jhc.4A6285.2004; Jacobberger JW, 2011, METHODS MOL BIOL, V699, P229, DOI 10.1007/978-1-61737-950-5_11; Juan G, 2004, CURR PROTOC CYTOM; Knudsen ES, 2010, CLIN CANCER RES, V16, P1094, DOI 10.1158/1078-0432.CCR-09-0787; Kotecha N, 2010, CURR PROTOC CYTOM, V10, P10, DOI [10.1002/0471142956.cy1017s53, DOI 10.1002/0471142956.CY1017S53]; MOORE MAS, 1974, BLOOD, V44, P1; Ornatsky O, 2006, J IMMUNOL METHODS, V308, P68, DOI 10.1016/j.jim.2005.09.020; Ornatsky OI, 2008, ANAL CHEM, V80, P2539, DOI 10.1021/ac702128m; Ornatsky OI, 2008, J ANAL ATOM SPECTROM, V23, P463, DOI 10.1039/b710510j; Passegue E, 2005, J EXP MED, V202, P1599, DOI 10.1084/jem.20050967; Pearce DJ, 2006, BLOOD, V107, P1166, DOI 10.1182/blood-2005-06-2325; PREISLER HD, 1980, BLOOD, V56, P361; Qiu P, 2011, NAT BIOTECHNOL, V29, P886, DOI 10.1038/nbt.1991; Raza A, 1997, EXP HEMATOL, V25, P530; Ren SJ, 2004, CELL, V117, P239, DOI 10.1016/S0092-8674(04)00300-9; Rothaeusler K, 2006, CYTOM PART A, V69A, P249, DOI 10.1002/cyto.a.20252; van Lochem EG, 2004, CYTOM PART B-CLIN CY, V60B, P1, DOI 10.1002/cyto.b.20008	32	114	114	3	48	WILEY	HOBOKEN	111 RIVER ST, HOBOKEN 07030-5774, NJ USA	1552-4922	1552-4930		CYTOM PART A	Cytom. Part A	JUL	2012	81A	7					552	566		10.1002/cyto.a.22075			15	Biochemical Research Methods; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	962PZ	WOS:000305558700006	22693166	Green Accepted			2020-07-01	
J	Fergusson, JR; Smith, KE; Fleming, VM; Rajoriya, N; Newell, EW; Simmons, R; Marchi, E; Bjorkander, S; Kang, YH; Swadling, L; Kurioka, A; Sahgal, N; Lockstone, H; Baban, D; Freeman, GJ; Sverremark-Ekstrom, E; Davis, MM; Davenport, MP; Venturi, V; Ussher, JE; Willberg, CB; Klenerman, P				Fergusson, Joannah R.; Smith, Kira E.; Fleming, Vicki M.; Rajoriya, Neil; Newell, Evan W.; Simmons, Ruth; Marchi, Emanuele; Bjorkander, Sophia; Kang, Yu-Hoi; Swadling, Leo; Kurioka, Ayako; Sahgal, Natasha; Lockstone, Helen; Baban, Dilair; Freeman, Gordon J.; Sverremark-Ekstrom, Eva; Davis, Mark M.; Davenport, Miles P.; Venturi, Vanessa; Ussher, James E.; Willberg, Christian B.; Klenerman, Paul			CD161 Defines a Transcriptional and Functional Phenotype across Distinct Human T Cell Lineages	CELL REPORTS			English	Article							HUMAN NKR-P1A; CUTTING EDGE; EXPRESSION; MAIT; SUBSET; LIGAND; NK; ACTIVATION; PROGRAM; NKRP1A	The C-type lectin CD161 is expressed by a large proportion of human T lymphocytes of all lineages, including a population known as mucosal-associated invariant T (MAIT) cells. To understand whether different T cell subsets expressing CD161 have similar properties, we examined these populations in parallel using mass cytometry and mRNA microarray approaches. The analysis identified a conserved CD161++/MAIT cell transcriptional signature enriched in CD161+CD8+ T cells, which can be extended to CD161+ CD4+ and CD161+TCR gamma delta+ T cells. Furthermore, this led to the identification of a shared innate-like, TCR-independent response to interleukin (IL)-12 plus IL-18 by different CD161-expressing T cell populations. This response was independent of regulation by CD161, which acted as a costimulatory molecule in the context of T cell receptor stimulation. Expression of CD161 hence identifies a transcriptional and functional phenotype, shared across human T lymphocytes and independent of both T cell receptor (TCR) expression and cell lineage.	[Fergusson, Joannah R.; Smith, Kira E.; Fleming, Vicki M.; Rajoriya, Neil; Simmons, Ruth; Marchi, Emanuele; Kang, Yu-Hoi; Swadling, Leo; Kurioka, Ayako; Ussher, James E.; Willberg, Christian B.; Klenerman, Paul] Univ Oxford, Peter Medawar Bldg Pathogen Res, Oxford OX1 3SY, England; [Fleming, Vicki M.] Oxford Univ Hosp NHS Trust, Dept Microbiol & Infect Dis, Oxford OX3 9DU, England; [Newell, Evan W.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Newell, Evan W.] Singapore Immunol Network SIgN, Agcy Sci Technol & Res A STAR, Singapore 138632, Singapore; [Bjorkander, Sophia; Sverremark-Ekstrom, Eva] Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, S-10691 Stockholm, Sweden; [Sahgal, Natasha; Lockstone, Helen; Baban, Dilair] Univ Oxford, Wellcome Trust Ctr Human Genet, Bioinformat & Stat Genet Core, Oxford OX3 7BN, England; [Freeman, Gordon J.] Harvard Univ, Sch Med, Dana Farber Canc Inst, Boston, MA 02215 USA; [Davenport, Miles P.; Venturi, Vanessa] Prince Wales Hosp, Dept Hematol, Kensington, NSW 2052, Australia; [Ussher, James E.] Univ Otago, Dept Microbiol & Immunol, Dunedin 9054, New Zealand; [Klenerman, Paul] John Radcliffe Hosp, NIHR Oxford Biomed Res Ctr, Oxford OX3 9TU, England[Fergusson, Joannah R.; Smith, Kira E.; Fleming, Vicki M.; Rajoriya, Neil; Simmons, Ruth; Marchi, Emanuele; Kang, Yu-Hoi; Swadling, Leo; Kurioka, Ayako; Ussher, James E.; Willberg, Christian B.; Klenerman, Paul] Univ Oxford, Peter Medawar Bldg Pathogen Res, Oxford OX1 3SY, England; [Fleming, Vicki M.] Oxford Univ Hosp NHS Trust, Dept Microbiol & Infect Dis, Oxford OX3 9DU, England; [Newell, Evan W.; Davis, Mark M.] Stanford Univ, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Newell, Evan W.] Singapore Immunol Network SIgN, Agcy Sci Technol & Res A STAR, Singapore 138632, Singapore; [Bjorkander, Sophia; Sverremark-Ekstrom, Eva] Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, S-10691 Stockholm, Sweden; [Sahgal, Natasha; Lockstone, Helen; Baban, Dilair] Univ Oxford, Wellcome Trust Ctr Human Genet, Bioinformat & Stat Genet Core, Oxford OX3 7BN, England; [Freeman, Gordon J.] Harvard Univ, Sch Med, Dana Farber Canc Inst, Boston, MA 02215 USA; [Davenport, Miles P.; Venturi, Vanessa] Prince Wales Hosp, Dept Hematol, Kensington, NSW 2052, Australia; [Ussher, James E.] Univ Otago, Dept Microbiol & Immunol, Dunedin 9054, New Zealand; [Klenerman, Paul] John Radcliffe Hosp, NIHR Oxford Biomed Res Ctr, Oxford OX3 9TU, England	Klenerman, P (reprint author), Univ Oxford, Peter Medawar Bldg Pathogen Res, Oxford OX1 3SY, England.	paul.klenerman@ndm.ox.ac.ukpaul.klenerman@ndm.ox.ac.uk	Ussher, James/O-4493-2019; Newell, Evan W/F-9711-2012; Freeman, Gordon/AAC-5380-2019; Sverremark-Ekstrom, Eva/A-2305-2016; Newell, Evan/AAE-9470-2020; Swadling, Leo/Y-5141-2019	Ussher, James/0000-0001-9222-7680; Newell, Evan W/0000-0002-2889-243X; Freeman, Gordon/0000-0002-7210-5616; Sverremark-Ekstrom, Eva/0000-0001-6271-8681; Newell, Evan/0000-0002-2889-243X; Swadling, Leo/0000-0002-0537-6715; Davenport, Miles/0000-0002-4751-1831; klenerman, paul/0000-0003-4307-9161; Willberg, Christian/0000-0001-5299-9344; Fergusson, Joannah/0000-0001-9223-1209	Wellcome Trust IITM Programme [092871/Z/10/Z]; Wellcome TrustWellcome Trust [WT091663MA]; Medical Research CouncilMedical Research Council UK (MRC); NIHR Biomedical Research Centre (Oxford)National Institute for Health Research (NIHR); Nuffield Department of Clinical Medicine (Oxford); James Martin School for the 21st Century (Oxford); NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [NIAD U19AI 082630]; Oxford Dominions Trust; National Institute for Health ResearchNational Institute for Health Research (NIHR) [NF-SI-0510-10204]	The authors would like to thank T. Hansen for the kind provision of the MR1-blocking antibody, N. Rust for sorting expertise, C. Marchant for technical support, and all members of the laboratory for helpful discussion and support. J.R.F is supported by the Wellcome Trust IITM Programme (092871/Z/10/Z). This work was also supported by the Wellcome Trust (WT091663MA), the Medical Research Council, the NIHR Biomedical Research Centre (Oxford), the Nuffield Department of Clinical Medicine (Oxford), the James Martin School for the 21st Century (Oxford), the NIH (NIAD U19AI 082630), and the Oxford Dominions Trust.	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J	Fienberg, HG; Simonds, EF; Fantl, WJ; Nolan, GP; Bodenmiller, B				Fienberg, Harris G.; Simonds, Erin F.; Fantl, Wendy J.; Nolan, Garry P.; Bodenmiller, Bernd			A platinum-based covalent viability reagent for single-cell mass cytometry	CYTOMETRY PART A			English	Article						mass cytometry; cisplatin; viability reagent	FLOW-CYTOMETRY; ICP-MS; CISPLATIN; DNA; APOPTOSIS; PHOSPHORYLATION; IDENTIFICATION; SPECTROMETRY; IMMUNOASSAY; SURFACE	In fluorescence-based flow cytometry, cellular viability is determined with membrane-impermeable fluorescent reagents that specifically enter and label plasma membrane-compromised nonviable cells. A recent technological advance in flow cytometry uses antibodies conjugated to elemental metal isotopes, rather than to fluorophores, to allow signal detection by atomic mass spectrometry. Unhampered by the limitations of overlapping emission fluorescence, mass cytometry increases the number of parameters that can be measured in single cells. However, mass cytometry is unable to take advantage of current fluorescent viability dyes. An alternative methodology was therefore developed here in which the platinum-containing chemotherapy drug cisplatin was used to resolve live and dead cells by mass cytometry. In a 1-min incubation step, cisplatin preferentially labeled nonviable cells from both adherent and suspension cultures, resulting in a platinum signal quantifiable by mass cytometry. This protocol was compatible with established sample processing steps for intracellular cytometry. Furthermore, the live/dead ratios were comparable between mass- and fluorescence-based cytometry. Importantly, although cisplatin is a known DNA-damaging agent, a 1-min pulse of cisplatin did not induce observable DNA damage or apoptotic responses even within 6-h post-exposure. Cisplatin can therefore be used as a viability reagent for a wide range of mass cytometry protocols. (c) 2012 International Society for Advancement of Cytometry	[Fienberg, Harris G.; Simonds, Erin F.; Fantl, Wendy J.; Nolan, Garry P.; Bodenmiller, Bernd] Stanford Univ, Sch Med, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Fienberg, Harris G.; Simonds, Erin F.; Fantl, Wendy J.; Nolan, Garry P.; Bodenmiller, Bernd] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA[Fienberg, Harris G.; Simonds, Erin F.; Fantl, Wendy J.; Nolan, Garry P.; Bodenmiller, Bernd] Stanford Univ, Sch Med, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Fienberg, Harris G.; Simonds, Erin F.; Fantl, Wendy J.; Nolan, Garry P.; Bodenmiller, Bernd] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA	Nolan, GP (reprint author), 269 Campus Dr,CCSR 3205, Stanford, CA 94305 USA.	gnolan@stanford.edu; bernd.bodenmiller@imls.uzh.chgnolan@stanford.edu; bernd.bodenmiller@imls.uzh.ch	Nolan, Garry/AAE-7903-2019	Simonds, Erin/0000-0002-3497-4861	Bill and Melinda Gates FoundationGates Foundation [GF12141-137101]; Lucille P Markey Stanford Graduate Fellowship; Swiss National Science FoundationSwiss National Science Foundation (SNSF); European Molecular Biology OrganizationEuropean Molecular Biology Organization (EMBO); Marie Curie IOFEuropean Union (EU); Rachford and Carlota A. Harris Endowed Professorship;  [P01 CA034233 (NCI)];  [1R01CA130826 (NCI)];  [RFA CA 09-011 (NCI)];  [U54CA149145 (NCI)];  [5U54CA143907 (NCI)];  [U19 AI057229 (NIAID)];  [HHSN272200700038C (NIAID)];  [DR1-01477 (CIRM)];  [RB2-01592 (CIRM)];  [NHLBI-HV-10-05(2) (NHLBI)];  [HEALTH.2010.1.2-1 (European Commission)]	Grant numbers: P01 CA034233 (NCI), 1R01CA130826 (NCI), RFA CA 09-011 (NCI), U54CA149145 (NCI), 5U54CA143907 (NCI), U19 AI057229 (NIAID), HHSN272200700038C (NIAID), DR1-01477 (CIRM), RB2-01592 (CIRM), NHLBI-HV-10-05(2) (NHLBI), HEALTH.2010.1.2-1 (European Commission), and GF12141-137101 (Bill and Melinda Gates Foundation). Grant sponsors: Lucille P Markey Stanford Graduate Fellowship (to HGF), Rachford and Carlota A. Harris Endowed Professorship (to GPN), Swiss National Science Foundation (to BB), the European Molecular Biology Organization (to BB), and the Marie Curie IOF (to BB).	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Part A	JUN	2012	81A	6					467	475		10.1002/cyto.a.22067			9	Biochemical Research Methods; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	945FO	WOS:000304258400008	22577098	Green Accepted			2020-07-01	
J	Bjornson, ZB; Nolan, GP; Fantl, WJ				Bjornson, Zach B.; Nolan, Garry P.; Fantl, Wendy J.			Single-cell mass cytometry for analysis of immune system functional states	CURRENT OPINION IN IMMUNOLOGY			English	Review							POLYCHROMATIC FLOW-CYTOMETRY; ELEMENT-TAGGED IMMUNOASSAY; REGULATORY T-CELLS; DIFFERENTIAL IMMUNE; VIRUS-INFECTION; SOLID TUMOR; ICP-MS; CYCLE; RESPONSES; INFLAMMATION	Mass cytometry facilitates high-dimensional, quantitative analysis of the effects of bioactive molecules on cell populations at single-cell resolution. Datasets are generated with panels of up to 45 antibodies. Each antibody is conjugated to a polymer chelated with a stable metal isotope, usually in the lanthanide series of the periodic table. Antibody panels recognize surface markers to delineate cell types simultaneously with intracellular signaling molecules to measure biological functions, such as metabolism, survival, DNA damage, cell cycle and apoptosis, to provide an overall determination of the network state of an individual cell. This review will cover the basics of mass cytometry as well as outline assays developed for the platform that enhance the immunologist's analytical arsenal.	[Bjornson, Zach B.; Nolan, Garry P.; Fantl, Wendy J.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA[Bjornson, Zach B.; Nolan, Garry P.; Fantl, Wendy J.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ, Sch Med, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, 269 Campus Dr, Stanford, CA 94305 USA.	gnolan@stanford.edu; wjfantl@stanford.edugnolan@stanford.edu; wjfantl@stanford.edu	Nolan, Garry/AAE-7903-2019		Rachford and Carlota A. Harris Endowed Chair; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, R01CA130826, U54CA149145, N01-HV-00242, 5-24927, 5U54CA143907]; FDAUnited States Department of Health & Human Services [HHSF223201210194C]; CIRMCalifornia Institute for Regenerative Medicine [DR1-01477, RB2-01592]; European Commission grantEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; DoD CDMRP Teal Innovator Award	The authors wish to thank Drs Scott Tanner, Olga Omatsky, Dmitry Bandura, Mitch Winnik and Mark Nitz for their critical reading of this manuscript. This work was supported by the Rachford and Carlota A. Harris Endowed Chair to GPN as well as NIH grants U19 AI057229, 1R01CA130826, U54CA149145, N01-HV-00242, 5-24927, 5U54CA143907, FDA contract HHSF223201210194C, CIRM grants DR1-01477 and RB2-01592, European Commission grant HEALTH.2010.1.2-1 and a DoD CDMRP Teal Innovator Award. Conflict of interest statement: G.P.N. has personal financial interest in the company DVS Sciences, the manufacturers of the instrument and reagents described in this manuscript.	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van der Maaten L., 2009, P 12 INT C ART INT S, P384; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; van Lochem EG, 2004, CYTOM PART B-CLIN CY, V60B, P1, DOI 10.1002/cyto.b.20008; Wali A, 1999, DNA CELL BIOL, V18, P837, DOI 10.1089/104454999314836; Yoo NK, 2008, CELL MICROBIOL, V10, P583, DOI 10.1111/j.1462-5822.2007.01047.x; Zhang YW, 2006, CELL CYCLE, V5, P125, DOI 10.4161/cc.5.2.2308	94	103	105	12	125	CURRENT BIOLOGY LTD	LONDON	84 THEOBALDS RD, LONDON WC1X 8RR, ENGLAND	0952-7915	1879-0372		CURR OPIN IMMUNOL	Curr. Opin. Immunol.	AUG	2013	25	4					484	494		10.1016/j.coi.2013.07.004			11	Immunology	Immunology	229BT	WOS:000325237500010	23999316	Green Accepted			2020-07-01	
J	Wu, MY; Singh, AK				Wu, Meiye; Singh, Anup K.			Single-cell protein analysis	CURRENT OPINION IN BIOTECHNOLOGY			English	Review							FLOW-CYTOMETRY; MASS CYTOMETRY; IDENTIFICATION; TRANSCRIPTOME; ENUMERATION; NETWORKS; DYNAMICS; ELISPOT; IMMUNE	Heterogeneity of cellular systems has been widely recognized but only recently have tools become available that allow probing of genes and proteins in single cells to understand it. While the advancement in single cell genomic analysis has been greatly aided by the power of amplification techniques (e.g. PCR), analysis of proteins in single cells has proven to be more challenging. However, recent advances in multi-parameter flow cytometry, microscopy, microfluidics and other techniques have made it possible to measure wide variety of proteins in single cells. In this review, we highlight key recent developments in analysis of proteins in a single cell (excluding imaging-based methods), and discuss their significance in biological research.	[Wu, Meiye; Singh, Anup K.] Sandia Natl Labs, Dept Bioengn & Biotechnol, Livermore, CA 94550 USA[Wu, Meiye; Singh, Anup K.] Sandia Natl Labs, Dept Bioengn & Biotechnol, Livermore, CA 94550 USA	Singh, AK (reprint author), Sandia Natl Labs, Dept Bioengn & Biotechnol, Livermore, CA 94550 USA.	aksingh@sandia.govaksingh@sandia.gov		Wu, Meiye/0000-0003-3712-1554	NIDCRUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Dental & Craniofacial Research (NIDCR) [R01 DE020891, P50GM085273]; NIGMSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of General Medical Sciences (NIGMS); ENIGMA, a Lawrence Berkeley National LaboratoryEuropean Union (EU); U.S. Department of Energy, Office of Science, Office of Biological and Environmental ResearchUnited States Department of Energy (DOE); United States Department of EnergyUnited States Department of Energy (DOE) [DE-AC04-94AL85000]	Financial support for preparation and some of the work included was provided by the grants: R01 DE020891, funded by the NIDCR; P50GM085273 (the New Mexico Spatiotemporal Modeling Center) funded by the NIGMS; and ENIGMA, a Lawrence Berkeley National Laboratory Scientific Focus Area Program supported by the U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research. Sandia is a multi-program laboratory operated by Sandia Corp., a Lockheed Martin Co., for the United States Department of Energy under Contract DE-AC04-94AL85000.	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Opin. Biotechnol.	FEB	2012	23	1					83	88		10.1016/j.copbio.2011.11.023			6	Biochemical Research Methods; Biotechnology & Applied Microbiology	Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology	900DZ	WOS:000300868100014	22189001	Green Accepted			2020-07-01	
J	Weber, LM; Robinson, MD				Weber, Lukas M.; Robinson, Mark D.			Comparison of Clustering Methods for High-Dimensional Single-Cell Flow and Mass Cytometry Data	CYTOMETRY PART A			English	Article						flow cytometry; mass cytometry; CyTOF; bioinformatics; clustering; manual gating; F1 score; high-dimensional; single-cell; cell populations	AUTOMATED IDENTIFICATION; POPULATIONS; BIOCONDUCTOR; SUBSETS; REVEALS; IMMUNE	Recent technological developments in high-dimensional flow cytometry and mass cytometry (CyTOF) have made it possible to detect expression levels of dozens of protein markers in thousands of cells per second, allowing cell populations to be characterized in unprecedented detail. Traditional data analysis by "manual gating" can be inefficient and unreliable in these high-dimensional settings, which has led to the development of a large number of automated analysis methods. Methods designed for unsupervised analysis use specialized clustering algorithms to detect and define cell populations for further downstream analysis. Here, we have performed an up-to-date, extensible performance comparison of clustering methods for high-dimensional flow and mass cytometry data. We evaluated methods using several publicly available data sets from experiments in immunology, containing both major and rare cell populations, with cell population identities from expert manual gating as the reference standard. Several methods performed well, including FlowSOM, X-shift, PhenoGraph, Rclusterpp, and flowMeans. Among these, FlowSOM had extremely fast runtimes, making this method well-suited for interactive, exploratory analysis of large, high-dimensional data sets on a standard laptop or desktop computer. These results extend previously published comparisons by focusing on high-dimensional data and including new methods developed for CyTOF data. R scripts to reproduce all analyses are available from GitHub (https://github.com/lmweber/cytometry-clustering-comparison), and pre-processed data files are available from Flow Repository (FR-FCM-ZZPH), allowing our comparisons to be extended to include new clustering methods and reference data sets. (C) 2016 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of ISAC.	[Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, SIB Swiss Inst Bioinformat, Zurich, Switzerland[Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland; [Weber, Lukas M.; Robinson, Mark D.] Univ Zurich, SIB Swiss Inst Bioinformat, Zurich, Switzerland	Robinson, MD (reprint author), Univ Zurich, Inst Mol Life Sci, Zurich, Switzerland.	mark.robinson@imls.uzh.chmark.robinson@imls.uzh.ch	Robinson, Mark/A-6432-2015	Robinson, Mark/0000-0002-3048-5518; Weber, Lukas M./0000-0002-3282-1730			Aghaeepour N, 2016, CYTOM PART A, V89A, P16, DOI 10.1002/cyto.a.22732; Aghaeepour N, 2013, NAT METHODS, V10, P228, DOI [10.1038/NMETH.2365, 10.1038/nmeth.2365]; Aghaeepour N, 2011, CYTOM PART A, V79A, P6, DOI 10.1002/cyto.a.21007; Becher B, 2014, NAT IMMUNOL, V15, P1181, DOI 10.1038/ni.3006; Bendall SC, 2012, TRENDS IMMUNOL, V33, P323, DOI 10.1016/j.it.2012.02.010; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Chen H, 2016, PLOS COMPUT BIOL, V12, DOI 10.1371/journal.pcbi.1005112; Chester C, 2015, J IMMUNOL, V195, P773, DOI 10.4049/jimmunol.1500633; Diggins KE, 2015, METHODS, V82, P55, DOI 10.1016/j.ymeth.2015.05.008; Finak G, 2016, SCI REP-UK, V6, DOI 10.1038/srep20686; Finak G, 2014, PLOS COMPUT BIOL, V10, DOI 10.1371/journal.pcbi.1003806; Finak Greg, 2009, Advances in Bioinformatics, V2009, P247646, DOI 10.1155/2009/247646; Ge Y, 2012, BIOINFORMATICS, V28, P2052, DOI 10.1093/bioinformatics/bts300; Hartmann FJ, 2016, J EXP MED, V213, P2621, DOI 10.1084/jem.20160897; Hornik K., 2005, J STAT SOFTW, P14; Huber W, 2015, NAT METHODS, V12, P115, DOI [10.1038/nmeth.3252, 10.1038/NMETH.3252]; Kriegel HP, 2009, ACM T KNOWL DISCOV D, V3, DOI 10.1145/1497577.1497578; Levine JH, 2015, CELL, V162, P184, DOI 10.1016/j.cell.2015.05.047; Linderman M., 2013, RCLUSTERPP LINKABLE; Lo K, 2009, BMC BIOINFORMATICS, V10, DOI 10.1186/1471-2105-10-145; Mair F, 2016, EUR J IMMUNOL, V46, P34, DOI 10.1002/eji.201545774; Mosmann TR, 2014, CYTOM PART A, V85, P422, DOI 10.1002/cyto.a.22445; Newell EW, 2016, NAT IMMUNOL, V17, P890, DOI 10.1038/ni.3485; Nilsson AR, 2013, CYTOM PART A, V83A, P721, DOI [10.1002/cyto.22324, 10.1002/cyto.a.22324]; Pejoski D, 2016, J IMMUNOL, V196, P4814, DOI 10.4049/jimmunol.1502005; Qian Y, 2010, CYTOM PART B-CLIN CY, V78B, pS69, DOI 10.1002/cyto.b.20554; Qiu P, 2011, NAT BIOTECHNOL, V29, P886, DOI 10.1038/nbt.1991; Ronan T, 2016, SCI SIGNAL, V9, DOI 10.1126/scisignal.aad1932; Saeys Y, 2016, NAT REV IMMUNOL, V1, P14; Samusik N, 2016, NAT METHODS, P1; Shekhar K, 2014, P NATL ACAD SCI USA, V111, P202, DOI 10.1073/pnas.1321405111; Sorensen T, 2015, CYTOM PART A, V87A, P603, DOI 10.1002/cyto.a.22626; Spidlen J, 2012, CYTOM PART A, V81A, P727, DOI 10.1002/cyto.a.22106; Van Gassen S, 2015, CYTOM PART A, V87A, P636, DOI 10.1002/cyto.a.22625; van Unen V, 2016, IMMUNITY, V44, P1227, DOI 10.1016/j.immuni.2016.04.014; Wiwie C, 2015, NAT METHODS, V12, P1033, DOI [10.1038/NMETH.3583, 10.1038/nmeth.3583]; Zare H, 2010, BMC BIOINFORMATICS, V11, DOI 10.1186/1471-2105-11-403	37	100	100	2	21	WILEY-BLACKWELL	HOBOKEN	111 RIVER ST, HOBOKEN 07030-5774, NJ USA	1552-4922	1552-4930		CYTOM PART A	Cytom. Part A	DEC	2016	89A	12					1084	1096		10.1002/cyto.a.23030			13	Biochemical Research Methods; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	EI7ZU	WOS:000392724500006	27992111	Green Accepted, Other Gold			2020-07-01	
J	Zunder, ER; Lujan, E; Goltsev, Y; Wernig, M; Nolan, GP				Zunder, Eli R.; Lujan, Ernesto; Goltsev, Yury; Wernig, Marius; Nolan, Garry P.			A Continuous Molecular Roadmap to iPSC Reprogramming through Progression Analysis of Single-Cell Mass Cytometry	CELL STEM CELL			English	Article							PLURIPOTENT STEM-CELLS; SOMATIC-CELLS; SELF-RENEWAL; SURFACE MARKERS; C-MYC; EXPRESSION; MOUSE; PATHWAY; MURINE; DIFFERENTIATION	To analyze cellular reprogramming at the single-cell level, mass cytometry was used to simultaneously measure markers of pluripotency, differentiation, cell-cycle status, and cellular signaling throughout the reprogramming process. Time-resolved progression analysis of the resulting data sets was used to construct a continuous molecular roadmap for three independent reprogramming systems. Although these systems varied substantially in Oct4, Sox2, Klf4, and c-Myc stoichiometry, they presented a common set of reprogramming landmarks. Early in the reprogramming process, Oct4(high) Klf4(high) cells transitioned to a CD73(high) CD104(high) CD54(low) partially reprogrammed state. Ki67(low) cells from this intermediate population reverted to a MEF-like phenotype, but Ki67(high) cells advanced through the M-E-T and then bifurcated into two distinct populations: an ESC-like Nanog(high) Sox2(high) CD54(high) population and a mesendoderm-like Nanog low Sox2 low Lin28 high CD24(high) PDGFR-alpha(high) population. The methods developed here for time-resolved, single-cell progression analysis may be used for the study of additional complex and dynamic systems, such as cancer progression and embryonic development.	[Zunder, Eli R.; Goltsev, Yury; Nolan, Garry P.] Stanford Univ Sch Med, Baxter Lab Stem Cell Biol, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Lujan, Ernesto; Wernig, Marius] Stanford Univ Sch Med, Dept Pathol, Inst Stem Cell Biol & Regenerat Med, Stanford, CA 94305 USA; [Lujan, Ernesto] Stanford Univ Sch Med, Dept Genet, Stanford, CA 94305 USA[Zunder, Eli R.; Goltsev, Yury; Nolan, Garry P.] Stanford Univ Sch Med, Baxter Lab Stem Cell Biol, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Lujan, Ernesto; Wernig, Marius] Stanford Univ Sch Med, Dept Pathol, Inst Stem Cell Biol & Regenerat Med, Stanford, CA 94305 USA; [Lujan, Ernesto] Stanford Univ Sch Med, Dept Genet, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ Sch Med, Baxter Lab Stem Cell Biol, Dept Microbiol & Immunol, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019		NIH NRSAUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [F32 GM093508-01]; California Institute for Regenerative Medicine (CIRM)California Institute for Regenerative Medicine [TG2-01159]; National Science Foundation (NSF) Graduate Research FellowshipNational Science Foundation (NSF) [DGE-114747]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19 AI057229, U54CA149145, N01-HV-00242, 1U19AI100627, 5R01AI07372405, R01CA184968, 1 R33 CA183654, R33 CA183692]; NIH-Baylor Research Institute [41000411217]; NIH-Northrop Grumman [7500108142]; CIRMCalifornia Institute for Regenerative Medicine [DR1-01477, RB2-01592]; Department of DefenseUnited States Department of Defense [OC110674, 11491122]; European CommissionEuropean Commission Joint Research Centre [Health.2010.1.2-1]; Food and Drug AdministrationUnited States Department of Health & Human Services [HHSF223201210194C]; Bill and Melinda Gates FoundationGates Foundation [OPP 1017093]; Alliance for Lupus Research; Entertainment Industry Foundation (NWCRA); Rachford and Carlota A. Harris Endowed Professorship	We thank Angelica Trejo and Astraea Jager for mass cytometry quality control and instrument maintenance. We thank the Stanford Shared FACS Facility for assistance with cell sorting. We thank the Stanford PAN Facility for assistance with microarray hybridization and imaging. We thank Rob Bruggner for patiently answering our questions about the R programming language. We thank Sean Bendall and David Burns for helpful discussions. We thank Alex Chang for assistance with immunofluorescence microscopy. E.R.Z. was supported by NIH NRSA F32 GM093508-01. E.L. was supported by the California Institute for Regenerative Medicine (CIRM) Predoctoral Fellowship TG2-01159 and the National Science Foundation (NSF) Graduate Research Fellowship DGE-114747. M.W. is a New York Stem Cell Foundation-Robertson Investigator and a Tashia and John Morgridge Faculty Scholar at the Child Health Research Institute at Stanford. This work was supported by NIH (U19 AI057229, U54CA149145, N01-HV-00242, 1U19AI100627, 5R01AI07372405, R01CA184968, 1 R33 CA183654, and R33 CA183692), NIH-Baylor Research Institute (41000411217), NIH-Northrop Grumman (7500108142), CIRM (DR1-01477 and RB2-01592), Department of Defense (OC110674 and 11491122), European Commission (Health.2010.1.2-1), Food and Drug Administration (HHSF223201210194C), Bill and Melinda Gates Foundation (OPP 1017093), the Alliance for Lupus Research, the Entertainment Industry Foundation (NWCRA grant), and the Rachford and Carlota A. Harris Endowed Professorship to G.P.N. G.P.N. has personal financial interest in the company Fluidigm, the manufacturer of the mass cytometer used in this manuscript.	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J	Bentzen, AK; Marquard, AM; Lyngaa, R; Saini, SK; Ramskov, S; Donia, M; Such, L; Furness, AJS; McGranahan, N; Rosenthal, R; Straten, PT; Szallasi, Z; Svane, IM; Swanton, C; Quezada, SA; Jakobsen, SN; Eklund, AC; Hadrup, SR				Bentzen, Amalie Kai; Marquard, Andrea Marion; Lyngaa, Rikke; Saini, Sunil Kumar; Ramskov, Sofie; Donia, Marco; Such, Lina; Furness, Andrew J. S.; McGranahan, Nicholas; Rosenthal, Rachel; Straten, Per Thor; Szallasi, Zoltan; Svane, Inge Marie; Swanton, Charles; Quezada, Sergio A.; Jakobsen, Soren Nyboe; Eklund, Aron Charles; Hadrup, Sine Reker			Large-scale detection of antigen-specific T cells using peptide-MHC-I multimers labeled with DNA barcodes	NATURE BIOTECHNOLOGY			English	Article							PARALLEL DETECTION; MASS CYTOMETRY; SELF-ANTIGEN; RESPONSES; MELANOMA; GENERATION; DESIGN; SENSITIVITY; RECOGNIZES; BLOCKADE	Identification of the peptides recognized by individual T cells is important for understanding and treating immune-related diseases. Current cytometry-based approaches are limited to the simultaneous screening of 10-100 distinct T-cell specificities in one sample. Here we use peptide-major histocompatibility complex (MHC) multimers labeled with individual DNA barcodes to screen >1,000 peptide specificities in a single sample, and detect low-frequency CD8 T cells specific for virus- or cancer restricted antigens. When analyzing T-cell recognition of shared melanoma antigens before and after adoptive cell therapy in melanoma patients, we observe a greater number of melanoma-specific T-cell populations compared with cytometry-based approaches. Furthermore, we detect neoepitope-specific T cells in tumor-infiltrating lymphocytes and peripheral blood from patients with non-small cell lung cancer. Barcode-labeled pMHC multimers enable the combination of functional T-cell analysis with large-scale epitope recognition profiling for the characterization of T-cell recognition in various diseases, including in small clinical samples.	[Bentzen, Amalie Kai; Lyngaa, Rikke; Saini, Sunil Kumar; Ramskov, Sofie; Such, Lina; Jakobsen, Soren Nyboe; Hadrup, Sine Reker] Tech Univ Denmark, Natl Vet Inst, Sect Immunol & Vaccinol, Copenhagen, Denmark; [Marquard, Andrea Marion; Szallasi, Zoltan; Eklund, Aron Charles] Tech Univ Denmark, Dept Syst Biol, Ctr Biol Sequence Anal, Lyngby, Denmark; [Donia, Marco; Straten, Per Thor; Svane, Inge Marie] Univ Copenhagen, Herlev Hosp, Dept Hematol, Ctr Canc Immune Therapy, Copenhagen, Denmark; [Donia, Marco; Svane, Inge Marie] Univ Copenhagen, Herlev Hosp, Dept Oncol, Copenhagen, Denmark; [Furness, Andrew J. S.; McGranahan, Nicholas; Rosenthal, Rachel; Swanton, Charles; Quezada, Sergio A.] UCL Canc Inst, CRUK Lung Canc Ctr Excellence, London, England; [Furness, Andrew J. S.; Quezada, Sergio A.] UCL, UCL Canc Inst, Canc Immunol Unit, London, England; [McGranahan, Nicholas; Rosenthal, Rachel; Swanton, Charles] Francis Crick Inst, Translat Canc Therapeut Lab, London, England; [Straten, Per Thor] Univ Copenhagen, Dept Immunol & Microbiol, Copenhagen, Denmark; [Jakobsen, Soren Nyboe] Immudex, Copenhagen, Denmark[Bentzen, Amalie Kai; Lyngaa, Rikke; Saini, Sunil Kumar; Ramskov, Sofie; Such, Lina; Jakobsen, Soren Nyboe; Hadrup, Sine Reker] Tech Univ Denmark, Natl Vet Inst, Sect Immunol & Vaccinol, Copenhagen, Denmark; [Marquard, Andrea Marion; Szallasi, Zoltan; Eklund, Aron Charles] Tech Univ Denmark, Dept Syst Biol, Ctr Biol Sequence Anal, Lyngby, Denmark; [Donia, Marco; Straten, Per Thor; Svane, Inge Marie] Univ Copenhagen, Herlev Hosp, Dept Hematol, Ctr Canc Immune Therapy, Copenhagen, Denmark; [Donia, Marco; Svane, Inge Marie] Univ Copenhagen, Herlev Hosp, Dept Oncol, Copenhagen, Denmark; [Furness, Andrew J. S.; McGranahan, Nicholas; Rosenthal, Rachel; Swanton, Charles; Quezada, Sergio A.] UCL Canc Inst, CRUK Lung Canc Ctr Excellence, London, England; [Furness, Andrew J. S.; Quezada, Sergio A.] UCL, UCL Canc Inst, Canc Immunol Unit, London, England; [McGranahan, Nicholas; Rosenthal, Rachel; Swanton, Charles] Francis Crick Inst, Translat Canc Therapeut Lab, London, England; [Straten, Per Thor] Univ Copenhagen, Dept Immunol & Microbiol, Copenhagen, Denmark; [Jakobsen, Soren Nyboe] Immudex, Copenhagen, Denmark	Hadrup, SR (reprint author), Tech Univ Denmark, Natl Vet Inst, Sect Immunol & Vaccinol, Copenhagen, Denmark.	sirha@vet.dtu.dksirha@vet.dtu.dk	Hadrup, Sine Reker/P-3388-2014	Hadrup, Sine Reker/0000-0002-5937-4344; Quezada, Sergio/0000-0002-9763-1700; Marquard, Andrea/0000-0003-2928-6017; Saini, Sunil Kumar/0000-0003-3382-9432; Svane, Inge Marie/0000-0002-9451-6037; Szallasi, Zoltan/0000-0001-5395-7509; Ramskov, Sofie/0000-0001-9413-8673; Donia, Marco/0000-0003-4966-9752; jakobsen, soren nyboe/0000-0002-2227-8615; thor Straten, Per/0000-0002-4731-4969; Swanton, Charles/0000-0002-4299-3018; McGranahan, Nicholas/0000-0001-9537-4045; Eklund, Aron Charles/0000-0003-0861-1001	Danish Cancer SocietyDanish Cancer Society [ID:R72-A4531-13-S2]; Lundbeck Foundation Fellowship [R190-2014-4178]; Danish Research Council (FSS)Det Frie Forskningsrad (DFF)Danish Medical Research Council [1331-00283]; Danish Research Council (DFF) [4004-00422]; Familien Erichsens Mindefond; Cancer Research UKCancer Research UK [FC001169]; UK Medical Research CouncilMedical Research Council UK (MRC) [FC001169, MR/FC001169/1]; Wellcome TrustWellcome Trust [FC001169]; Novo Nordisk FoundationNovo Nordisk Foundation [16584]; Cancer Research UKCancer Research UK [12100, 20265, 20466]; Lundbeck FoundationLundbeckfonden [R181-2014-3828]; Novo Nordisk FondenNovo NordiskNovo Nordisk Foundation [NNF15OC0016584]	We would like to thank U.K. Hansen, A. Burkal and T. Seremet for technical assistance; T. Schumacher, Netherlands Cancer Institute, for scientific discussions and sharing of MHC expression plasmids; and Dr. Altman, NIH Tetramer Core Facility, for sharing expression plasmids HLA-B*1501 and HLA-B*3501. The work was funded by The Danish Cancer Society (ID:R72-A4531-13-S2), The Lundbeck Foundation Fellowship (ID: R190-2014-4178), The Danish Research Council (FSS-ID: 1331-00283 and DFF-ID:4004-00422), Familien Erichsens Mindefond, Cancer Research UK (FC001169), the UK Medical Research Council (FC001169), the Wellcome Trust (FC001169), the UK Medical Research Council (MR/FC001169/1) and the Novo Nordisk Foundation (ID: 16584).	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Biotechnol.	OCT	2016	34	10					1037	1045		10.1038/nbt.3662			9	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	EA0ZG	WOS:000386317500018	27571370				2020-07-01	
J	Wong, MT; Ong, DEH; Lim, FSH; Teng, KWW; McGovern, N; Narayanan, S; Ho, WQ; Cerny, D; Tan, HKK; Anicete, R; Tan, BK; Lim, TKH; Chan, CY; Cheow, PC; Lee, SY; Takano, A; Tan, EH; Tam, JKC; Tan, EY; Chan, JKY; Fink, K; Bertoletti, A; Ginhoux, F; De Lafaille, MAC; Newell, EW				Wong, Michael Thomas; Ong, David Eng Hui; Lim, Frances Sheau Huei; Teng, Karen Wei Weng; McGovern, Naomi; Narayanan, Sriram; Ho, Wen Qi; Cerny, Daniela; Tan, Henry Kun Kiaang; Anicete, Rosslyn; Tan, Bien Keem; Lim, Tony Kiat Hon; Chan, Chung Yip; Cheow, Peng Chung; Lee, Ser Yee; Takano, Angela; Tan, Eng-Huat; Tam, John Kit Chung; Tan, Ern Yu; Chan, Jerry Kok Yen; Fink, Katja; Bertoletti, Antonio; Ginhoux, Florent; De Lafaille, Maria Alicia Curotto; Newell, Evan William			A High-Dimensional Atlas of Human T Cell Diversity Reveals Tissue-Specific Trafficking and Cytokine Signatures	IMMUNITY			English	Article							CHEMOKINE RECEPTOR EXPRESSION; MASS CYTOMETRY; INTERLEUKIN 22; TGF-BETA; DIFFERENTIATION; IMMUNITY; DISTINCT; SUBSET; COMPARTMENTALIZATION; LYMPHOCYTES	Depending on the tissue microenvironment, T cells can differentiate into highly diverse subsets expressing unique trafficking receptors and cytokines. Studies of human lymphocytes have primarily focused on a limited number of parameters in blood, representing an incomplete view of the human immune system. Here, we have utilized mass cytometry to simultaneously analyze T cell trafficking and functional markers across eight different human tissues, including blood, lymphoid, and non-lymphoid tissues. These data have revealed that combinatorial expression of trafficking receptors and cytokines better defines tissue specificity. Notably, we identified numerous T helper cell subsets with overlapping cytokine expression, but only specific cytokine combinations are secreted regardless of tissue type. This indicates that T cell lineages defined in mouse models cannot be clearly distinguished in humans. Overall, our data uncover a plethora of tissue immune signatures and provide a systemic map of how T cell phenotypes are altered throughout the human body.	[Wong, Michael Thomas; Teng, Karen Wei Weng; McGovern, Naomi; Narayanan, Sriram; Ho, Wen Qi; Cerny, Daniela; Fink, Katja; Bertoletti, Antonio; Ginhoux, Florent; De Lafaille, Maria Alicia Curotto; Newell, Evan William] Singapore Immunol Network SIgN, A STAR, Singapore 138648, Singapore; [Ong, David Eng Hui] Natl Univ Hlth Syst, Univ Med Cluster, Div Gastroenterol & Hepatol, Singapore 119228, Singapore; [Lim, Frances Sheau Huei] Natl Univ Hlth Syst, Univ Surg Ctr, Div Colorectal Surg, Singapore 119228, Singapore; [Cerny, Daniela] Nanyang Technol Univ, Sch Biol Sci, Singapore 637551, Singapore; [Tan, Henry Kun Kiaang; Anicete, Rosslyn] KK Womens & Childrens Hosp, Dept Otolaryngol, Singapore 229899, Singapore; [Tan, Bien Keem] Singapore Gen Hosp, Dept Plast Reconstruct & Aesthet Surg, Singapore 169608, Singapore; [Lim, Tony Kiat Hon; Takano, Angela] Singapore Gen Hosp, Dept Pathol, Singapore 169856, Singapore; [Chan, Chung Yip; Cheow, Peng Chung; Lee, Ser Yee] Singapore Gen Hosp, Dept Hepatopancreatobiliary & Transplant Surg, Singapore 169856, Singapore; [Tan, Eng-Huat] Natl Canc Ctr, Dept Med Oncol, Singapore 169610, Singapore; [Tam, John Kit Chung] Natl Univ Singapore, Dept Surg, Singapore 119228, Singapore; [Tan, Ern Yu] Tan Tock Seng Hosp, Dept Gen Surg, Singapore 308433, Singapore; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Singapore 229899, Singapore; [Bertoletti, Antonio] Duke NUS Med Sch, Emerging Infect Dis EID Program, Singapore 169857, Singapore[Wong, Michael Thomas; Teng, Karen Wei Weng; McGovern, Naomi; Narayanan, Sriram; Ho, Wen Qi; Cerny, Daniela; Fink, Katja; Bertoletti, Antonio; Ginhoux, Florent; De Lafaille, Maria Alicia Curotto; Newell, Evan William] Singapore Immunol Network SIgN, A STAR, Singapore 138648, Singapore; [Ong, David Eng Hui] Natl Univ Hlth Syst, Univ Med Cluster, Div Gastroenterol & Hepatol, Singapore 119228, Singapore; [Lim, Frances Sheau Huei] Natl Univ Hlth Syst, Univ Surg Ctr, Div Colorectal Surg, Singapore 119228, Singapore; [Cerny, Daniela] Nanyang Technol Univ, Sch Biol Sci, Singapore 637551, Singapore; [Tan, Henry Kun Kiaang; Anicete, Rosslyn] KK Womens & Childrens Hosp, Dept Otolaryngol, Singapore 229899, Singapore; [Tan, Bien Keem] Singapore Gen Hosp, Dept Plast Reconstruct & Aesthet Surg, Singapore 169608, Singapore; [Lim, Tony Kiat Hon; Takano, Angela] Singapore Gen Hosp, Dept Pathol, Singapore 169856, Singapore; [Chan, Chung Yip; Cheow, Peng Chung; Lee, Ser Yee] Singapore Gen Hosp, Dept Hepatopancreatobiliary & Transplant Surg, Singapore 169856, Singapore; [Tan, Eng-Huat] Natl Canc Ctr, Dept Med Oncol, Singapore 169610, Singapore; [Tam, John Kit Chung] Natl Univ Singapore, Dept Surg, Singapore 119228, Singapore; [Tan, Ern Yu] Tan Tock Seng Hosp, Dept Gen Surg, Singapore 308433, Singapore; [Chan, Jerry Kok Yen] KK Womens & Childrens Hosp, Dept Reprod Med, Singapore 229899, Singapore; [Bertoletti, Antonio] Duke NUS Med Sch, Emerging Infect Dis EID Program, Singapore 169857, Singapore	Wong, MT; Newell, EW (reprint author), Singapore Immunol Network SIgN, A STAR, Singapore 138648, Singapore.	michael.thomas.wong@merck.com; evan_newell@immunol.a-star.edu.sgmichael.thomas.wong@merck.com; evan_newell@immunol.a-star.edu.sg	Newell, Evan/AAE-9470-2020; Newell, Evan W/F-9711-2012; Lim, Frances/B-5168-2011; Tam, John/B-6048-2012	Newell, Evan/0000-0002-2889-243X; Newell, Evan W/0000-0002-2889-243X; McGovern, Naomi/0000-0001-5200-2698	A-STAR/SIgN core funding; A-STAR/SIgN immunomonitoring platform funding; Singapore Translational Research (STaR) Investigator Award [NMRC/STaR/013/2012]	The authors thank the Singapore Immunology Network (SIgN) community and all members of the Newell lab for helpful discussion and technical support. We thank the Clinical Trials Resource Centre (Singapore General Hospital) for providing select skin samples and the National Cancer Center Singapore (NCCS) Translational & Clinical Research (TCR) research coordinators for helping to provide select lung samples. We also thank Paul Klenerman and Laura Mackay for helpful comments. This study was primarily funded by A-STAR/SIgN core funding (E.W.N., F.G., M.A.C.D., and K.F.) and the A-STAR/SIgN immunomonitoring platform funding (E.W.N.) and was partially supported by a Singapore Translational Research (STaR) Investigator Award (NMRC/STaR/013/2012) to A. B.	Acosta-Rodriguez EV, 2007, NAT IMMUNOL, V8, P639, DOI 10.1038/ni1467; Amir ED, 2013, NAT BIOTECHNOL, V31, P545, DOI 10.1038/nbt.2594; Becher B, 2014, NAT IMMUNOL, V15, P1181, DOI 10.1038/ni.3006; Bendall SC, 2012, TRENDS IMMUNOL, V33, P323, DOI 10.1016/j.it.2012.02.010; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Campbell JJ, 2001, J IMMUNOL, V166, P6477, DOI 10.4049/jimmunol.166.11.6477; Cerny D, 2014, PLOS PATHOG, V10, DOI 10.1371/journal.ppat.1004548; Cheng Y, 2016, J IMMUNOL, V196, P924, DOI 10.4049/jimmunol.1501928; Cosmi L, 2008, J EXP MED, V205, P1903, DOI 10.1084/jem.20080397; Cox D, 2010, NAT REV DRUG DISCOV, V9, P804, DOI 10.1038/nrd3266; Duhen T, 2014, J IMMUNOL, V193, P120, DOI 10.4049/jimmunol.1302734; Duhen T, 2009, NAT IMMUNOL, V10, P857, DOI 10.1038/ni.1767; Fu HM, 2013, FRONT BIOSCI-LANDMRK, V18, P422, DOI 10.2741/4111; Ghoreschi K, 2010, NATURE, V467, P967, DOI 10.1038/nature09447; Gibbons D, 2014, NAT MED, V20, P1206, DOI 10.1038/nm.3670; Griffith JW, 2014, ANNU REV IMMUNOL, V32, P659, DOI 10.1146/annurev-immunol-032713-120145; Hamada H, 2009, J IMMUNOL, V182, P3469, DOI 10.4049/jimmunol.0801814; He DG, 2006, J IMMUNOL, V177, P6852, DOI 10.4049/jimmunol.177.10.6852; Hong M, 2015, NAT COMMUN, V6, DOI 10.1038/ncomms7588; Islam SA, 2012, NAT MED, V18, P705, DOI 10.1038/nm.2760; Kattah MG, 2008, ARTHRITIS RHEUM, V58, P1619, DOI 10.1002/art.23497; Kim CH, 2001, J CLIN INVEST, V108, P1331, DOI 10.1172/JCI200113543; Mackay LK, 2015, IMMUNITY, V43, P1101, DOI 10.1016/j.immuni.2015.11.008; Newell EW, 2014, NAT BIOTECHNOL, V32, P149, DOI 10.1038/nbt.2783; Newell EW, 2012, IMMUNITY, V36, P142, DOI 10.1016/j.immuni.2012.01.002; Park CO, 2015, NAT MED, V21, P688, DOI 10.1038/nm.3883; Sallusto F, 1998, J EXP MED, V187, P875, DOI 10.1084/jem.187.6.875; Sallusto F, 1999, NATURE, V401, P708, DOI 10.1038/44385; Sathaliyawala T, 2013, IMMUNITY, V38, P187, DOI 10.1016/j.immuni.2012.09.020; Shekhar K, 2014, P NATL ACAD SCI USA, V111, P202, DOI 10.1073/pnas.1321405111; Singh SP, 2008, J IMMUNOL, V180, P214, DOI 10.4049/jimmunol.180.1.214; Skon CN, 2013, NAT IMMUNOL, V14, P1285, DOI 10.1038/ni.2745; Tang XZ, 2013, J IMMUNOL, V190, P3142, DOI 10.4049/jimmunol.1203218; Thome JJC, 2016, NAT MED, V22, P72, DOI 10.1038/nm.4008; Thome JJC, 2015, TRENDS IMMUNOL, V36, P428, DOI 10.1016/j.it.2015.05.003; Thome JJC, 2014, CELL, V159, P814, DOI 10.1016/j.cell.2014.10.026; Trifari S, 2009, NAT IMMUNOL, V10, P864, DOI 10.1038/ni.1770; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; Viola A, 2008, ANNU REV PHARMACOL, V48, P171, DOI 10.1146/annurev.pharmtox.48.121806.154841; Wang J, 2014, J EXP MED, V211, P2397, DOI 10.1084/jem.20140625; WESTERMANN J, 1992, CLIN INVESTIGATOR, V70, P539; Wong MT, 2015, CELL REP, V11, P1822, DOI 10.1016/j.celrep.2015.05.022; Wong MT, 2010, IMMUNOL CELL BIOL, V88, P624, DOI 10.1038/icb.2010.53; Yamane H, 2013, IMMUNOL REV, V252, P12, DOI 10.1111/imr.12032; Yen HR, 2009, J IMMUNOL, V183, P7161, DOI 10.4049/jimmunol.0900368; Zhu JF, 2010, CELL RES, V20, P4, DOI 10.1038/cr.2009.138; Zuniga LA, 2013, IMMUNOL REV, V252, P78, DOI 10.1111/imr.12036	47	93	95	1	17	CELL PRESS	CAMBRIDGE	50 HAMPSHIRE ST, FLOOR 5, CAMBRIDGE, MA 02139 USA	1074-7613	1097-4180		IMMUNITY	Immunity	AUG 16	2016	45	2					442	456		10.1016/j.immuni.2016.07.007			15	Immunology	Immunology	DU5QV	WOS:000382267300023	27521270	Bronze			2020-07-01	
J	Krishnaswamy, S; Spitzer, MH; Mingueneau, M; Bendall, SC; Litvin, O; Stone, E; Pe'er, D; Nolan, GP				Krishnaswamy, Smita; Spitzer, Matthew H.; Mingueneau, Michael; Bendall, Sean C.; Litvin, Oren; Stone, Erica; Pe'er, Dana; Nolan, Garry P.			Conditional density-based analysis of T cell signaling in single-cell data	SCIENCE			English	Article							MASS CYTOMETRY; MUTUAL INFORMATION; CD4; ACTIVATION; GENERATION; RESPONSES; PATHWAYS; IMMUNE; NAIVE	Cellular circuits sense the environment, process signals, and compute decisions using networks of interacting proteins. To model such a system, the abundance of each activated protein species can be described as a stochastic function of the abundance of other proteins. High-dimensional single-cell technologies, such as mass cytometry, offer an opportunity to characterize signaling circuit-wide. However, the challenge of developing and applying computational approaches to interpret such complex data remains. Here, we developed computational methods, based on established statistical concepts, to characterize signaling network relationships by quantifying the strengths of network edges and deriving signaling response functions. In comparing signaling between naive and antigen-exposed CD4(+) T lymphocytes, we find that although these two cell subtypes had similarly wired networks, naive cells transmitted more information along a key signaling cascade than did antigen-exposed cells. We validated our characterization on mice lacking the extracellular-regulated mitogen-activated protein kinase (MAPK) ERK2, which showed stronger influence of pERK on pS6 (phosphorylated-ribosomal protein S6), in naive cells as compared with antigen-exposed cells, as predicted. We demonstrate that by using cell-to-cell variation inherent in single-cell data, we can derive response functions underlying molecular circuits and drive the understanding of how cells process signals.	[Krishnaswamy, Smita; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA; [Spitzer, Matthew H.; Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Mingueneau, Michael] Harvard Univ, Sch Med, Dept Microbiol & Immunobiol, Div Immunol, Boston, MA USA; [Stone, Erica] Univ Calif San Diego, Dept Cellular & Mol Med, Div Biol Sci, Mol Biol Sect, La Jolla, CA 92093 USA[Krishnaswamy, Smita; Litvin, Oren; Pe'er, Dana] Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA; [Spitzer, Matthew H.; Bendall, Sean C.; Nolan, Garry P.] Stanford Univ, Dept Microbiol & Immunol, Baxter Lab Stem Cell Biol, Stanford, CA 94305 USA; [Mingueneau, Michael] Harvard Univ, Sch Med, Dept Microbiol & Immunobiol, Div Immunol, Boston, MA USA; [Stone, Erica] Univ Calif San Diego, Dept Cellular & Mol Med, Div Biol Sci, Mol Biol Sect, La Jolla, CA 92093 USA	Pe'er, D (reprint author), Columbia Univ, Dept Biol Sci, Dept Syst Biol, New York, NY 10027 USA.	dpeer@biology.columbia.edudpeer@biology.columbia.edu	Krishnaswamy, Smita/D-8089-2016; Nolan, Garry/AAE-7903-2019; Mingueneau, Michael/O-5313-2019	Krishnaswamy, Smita/0000-0001-5823-1985; Mingueneau, Michael/0000-0002-3873-7329; Spitzer, Matthew/0000-0002-5291-3819	National Science Foundation CAREER awardNational Science Foundation (NSF) [MCB-1149728]; National Centers for Biomedical Computing Grant [1U54CA121852-01A1]; NIH S10 Shared instrumentation grant [NIH S10 SIG S10RR027582-01]; Packard Fellowship for Science and Engineering; Damon Runyon Cancer Research Foundation Fellowship [DRG-2017-09]; NIH K01 award [1K01DK095008]; Rachford and Carlota A. Harris Endowed Professorship; NCIUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [RFA CA 09-011, NHLBIHV-10-05 (2)]; European CommissionEuropean Commission Joint Research Centre [HEALTH.2010.1.2-1]; Bill and Melinda Gates FoundationGates Foundation [GF12141-137101];  [U19 AI057229];  [P01 CA034233];  [HHSN272200700038C];  [1R01CA130826];  [CIRM DR1-01477];  [RB2-01592]	We thank B. Bodenmiller, N. Friedman, N. HaCohen, I. Pe'er, A. Regev, and S. Reiner for valuable comments. We respectfully appreciate the provision of mouse samples and key mechanistic discussions with C. Benoist and S. Hedrick. This research was supported by the National Science Foundation CAREER award through grant MCB-1149728 and National Centers for Biomedical Computing Grant 1U54CA121852-01A1 and the NIH S10 Shared instrumentation grant NIH S10 SIG S10RR027582-01. D. P. holds a Packard Fellowship for Science and Engineering. S. C. B. is supported by the Damon Runyon Cancer Research Foundation Fellowship (DRG-2017-09). E. S. is supported by NIH K01 award 1K01DK095008. G. P. N. is supported by the Rachford and Carlota A. Harris Endowed Professorship and grants from U19 AI057229, P01 CA034233, HHSN272200700038C, 1R01CA130826, CIRM DR1-01477, and RB2-01592, NCI RFA CA 09-011, NHLBIHV-10-05 (2), European Commission HEALTH.2010.1.2-1, and the Bill and Melinda Gates Foundation (GF12141-137101). S. K., G. P. N., and D. P. conceived the study. S. K. and D. P. designed and developed DREVI, DREMI, and additional computational methods in this manuscript. S. K. wrote all computer programs used in this manuscript. M. H. S. and S. C. B. developed reagents. M. M. stimulated the mice and collected the biological samples. M. H. S., M. M., and S. C. B. designed mouse experiments and performed cognate data acquisition. E. S. developed the inducible ERK-knockdown mouse. S. K. and O.L. performed statistical analysis. S. K. and D. P. performed the biological analysis and interpretation. S. K., G. P. N., and D. P. wrote the manuscript.	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J	Mair, F; Hartmann, FJ; Mrdjen, D; Tosevski, V; Krieg, C; Becher, B				Mair, Florian; Hartmann, Felix J.; Mrdjen, Dunja; Tosevski, Vinko; Krieg, Carsten; Becher, Burkhard			The end of gating? An introduction to automated analysis of high dimensional cytometry data	EUROPEAN JOURNAL OF IMMUNOLOGY			English	Review						Citrus; CyTOF; Data analysis; Flow cytometry; Mass cytometry; PSM; PhenoGraph; SPADE; t-SNE; Wanderlust	HUMAN B-CELL; FLOW-CYTOMETRY; MASS CYTOMETRY; BIOLOGICAL CELLS; IDENTIFICATION; NORMALIZATION; POPULATIONS; PHENOTYPE; IMMUNE; GUIDE	Ever since its invention half a century ago, flow cytometry has been a major tool for single-cell analysis, fueling advances in our understanding of a variety of complex cellular systems, in particular the immune system. The last decade has witnessed significant technical improvements in available cytometry platforms, such that more than 20 parameters can be analyzed on a single-cell level by fluorescence-based flow cytometry. The advent of mass cytometry has pushed this limit up to, currently, 50 parameters. However, traditional analysis approaches for the resulting high-dimensional datasets, such as gating on bivariate dot plots, have proven to be inefficient. Although a variety of novel computational analysis approaches to interpret these datasets are already available, they have not yet made it into the mainstream and remain largely unknown to many immunologists. Therefore, this review aims at providing a practical overview of novel analysis techniques for high-dimensional cytometry data including SPADE, t-SNE, Wanderlust, Citrus, and PhenoGraph, and how these applications can be used advantageously not only for the most complex datasets, but also for standard 14-parameter cytometry datasets.	[Mair, Florian; Hartmann, Felix J.; Mrdjen, Dunja; Tosevski, Vinko; Krieg, Carsten; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, CH-8091 Zurich, Switzerland[Mair, Florian; Hartmann, Felix J.; Mrdjen, Dunja; Tosevski, Vinko; Krieg, Carsten; Becher, Burkhard] Univ Zurich, Inst Expt Immunol, CH-8091 Zurich, Switzerland	Becher, B (reprint author), Univ Zurich, Inst Expt Immunol, Sternwartstr 2, CH-8091 Zurich, Switzerland.	becher@immunology.uzh.chbecher@immunology.uzh.ch		Krieg, Carsten/0000-0002-5145-7591; Becher, Burkhard/0000-0002-1541-7867; Mrdjen, Dunja/0000-0001-9269-2806; Mair, Florian/0000-0001-6732-5449; Hartmann, Felix/0000-0002-4174-2276	Swiss National Science Foundation (SNF)Swiss National Science Foundation (SNSF) [316030_150768, 310030_146130, CRSII3_136203]; European Union FP7 project TargetBraIn; ATECT; University Research Priority Project: Translational cancer research; NeuroKine	This work was supported by grants from the Swiss National Science Foundation (SNF, accession numbers 316030_150768, 310030_146130, and CRSII3_136203), the European Union FP7 project TargetBraIn, NeuroKine, and ATECT and the University Research Priority Project: Translational cancer research.	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J. Immunol.	JAN	2016	46	1					34	43		10.1002/eji.201545774			10	Immunology	Immunology	DB0YO	WOS:000368234800005	26548301				2020-07-01	
J	Kidd, BA; Peters, LA; Schadt, EE; Dudley, JT				Kidd, Brian A.; Peters, Lauren A.; Schadt, Eric E.; Dudley, Joel T.			Unifying immunology with informatics and multiscale biology	NATURE IMMUNOLOGY			English	Review							DIFFERENTIAL EXPRESSION ANALYSIS; CHIP-SEQ EXPERIMENTS; HLA CLASS-I; GENE-EXPRESSION; SYSTEMS-BIOLOGY; HIGH-THROUGHPUT; REGULATORY NETWORKS; ENRICHMENT ANALYSIS; MASS CYTOMETRY; RNA-SEQ	The immune system is a highly complex and dynamic system. Historically, the most common scientific and clinical practice has been to evaluate its individual components. This kind of approach cannot always expose the interconnecting pathways that control immune-system responses and does not reveal how the immune system works across multiple biological systems and scales. High-throughput technologies can be used to measure thousands of parameters of the immune system at a genome-wide scale. These system-wide surveys yield massive amounts of quantitative data that provide a means to monitor and probe immune-system function. New integrative analyses can help synthesize and transform these data into valuable biological insight. Here we review some of the computational analysis tools for high-dimensional data and how they can be applied to immunology.	[Kidd, Brian A.; Schadt, Eric E.; Dudley, Joel T.] Dept Genet & Genom Sci, New York, NY 10029 USA; [Kidd, Brian A.; Schadt, Eric E.; Dudley, Joel T.] Icahn Inst Genom & Multiscale Biol, New York, NY USA; [Kidd, Brian A.; Peters, Lauren A.; Schadt, Eric E.; Dudley, Joel T.] Icahn Sch Med Mt Sinai, New York, NY USA; [Peters, Lauren A.] Grad Sch Biomed Sci, New York, NY USA[Kidd, Brian A.; Schadt, Eric E.; Dudley, Joel T.] Dept Genet & Genom Sci, New York, NY 10029 USA; [Kidd, Brian A.; Schadt, Eric E.; Dudley, Joel T.] Icahn Inst Genom & Multiscale Biol, New York, NY USA; [Kidd, Brian A.; Peters, Lauren A.; Schadt, Eric E.; Dudley, Joel T.] Icahn Sch Med Mt Sinai, New York, NY USA; [Peters, Lauren A.] Grad Sch Biomed Sci, New York, NY USA	Dudley, JT (reprint author), Dept Genet & Genom Sci, New York, NY 10029 USA.	joel.dudley@mssm.edujoel.dudley@mssm.edu		Kidd, Brian/0000-0003-2110-1145	National Institute of Diabetes and Digestive and Kidney DiseasesUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Diabetes & Digestive & Kidney Diseases (NIDDK) [R01 DK098242]; Pharmaceutical Research and Manufacturers of America Foundation	We thank C. Berin, B. Brown, R. Kosoy, B. Readhead and C. Tato for critical reading and feedback on the manuscript. This work was supported by funding from the National Institute of Diabetes and Digestive and Kidney Diseases (R01 DK098242) and the Pharmaceutical Research and Manufacturers of America Foundation.	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Immunol.	FEB	2014	15	2					118	127		10.1038/ni.2787			10	Immunology	Immunology	295XV	WOS:000330150600003	24448569	Green Accepted			2020-07-01	
J	Krijger, PHL; Di Stefano, B; de Wit, E; Limone, F; van Oevelen, C; de Laat, W; Graf, T				Krijger, Peter Hugo Lodewijk; Di Stefano, Bruno; de Wit, Elzo; Limone, Francesco; van Oevelen, Chris; de Laat, Wouter; Graf, Thomas			Cell-of-Origin-Specific 3D Genome Structure Acquired during Somatic Cell Reprogramming	CELL STEM CELL			English	Article							PLURIPOTENT STEM-CELLS; CHROMATIN ARCHITECTURE; EPIGENETIC MEMORY; MASS CYTOMETRY; IPS CELLS; DIFFERENTIATION; REORGANIZATION; ORGANIZATION; PRINCIPLES; ENHANCERS	Forced expression of reprogramming factors can convert somatic cells into induced pluripotent stem cells (iPSCs). Here we studied genome topology dynamics during reprogramming of different somatic cell types with highly distinct genome conformations. We find large-scale topologically associated domain (TAD) repositioning and alterations of tissue-restricted genomic neighborhoods and chromatin loops, effectively erasing the somatic-cell-specific genome structures while establishing an embryonic stem-cell-like 3D genome. Yet, early passage iPSCs carry topological hallmarks that enable recognition of their cell of origin. These hallmarks are not remnants of somatic chromosome topologies. Instead, the distinguishing topological features are acquired during reprogramming, as we also find for cell-of-origin-dependent gene expression patterns.	[Krijger, Peter Hugo Lodewijk; de Wit, Elzo; de Laat, Wouter] Hubrecht Inst KNAW, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands; [Krijger, Peter Hugo Lodewijk; de Wit, Elzo; de Laat, Wouter] Univ Med Ctr Utrecht, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands; [Di Stefano, Bruno; Limone, Francesco; van Oevelen, Chris; Graf, Thomas] Ctr Genom Regulat CRG, Gene Regulat Stem Cells & Canc Programme, Dr Aiguader 88, Barcelona 08003, Spain; [Di Stefano, Bruno; Limone, Francesco; van Oevelen, Chris; Graf, Thomas] Pompeu Fabra Univ, Dr Aiguader 88, Barcelona 08003, Spain; [Di Stefano, Bruno] Massachusetts Gen Hosp, Dept Mol Biol, Boston, MA 02114 USA; [Di Stefano, Bruno] Harvard Univ, Sch Med, Boston, MA 02114 USA; [de Wit, Elzo] Netherlands Canc Inst, Div Gene Regulat, Plesmanlaan 121, NL-1066 CX Amsterdam, Netherlands[Krijger, Peter Hugo Lodewijk; de Wit, Elzo; de Laat, Wouter] Hubrecht Inst KNAW, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands; [Krijger, Peter Hugo Lodewijk; de Wit, Elzo; de Laat, Wouter] Univ Med Ctr Utrecht, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands; [Di Stefano, Bruno; Limone, Francesco; van Oevelen, Chris; Graf, Thomas] Ctr Genom Regulat CRG, Gene Regulat Stem Cells & Canc Programme, Dr Aiguader 88, Barcelona 08003, Spain; [Di Stefano, Bruno; Limone, Francesco; van Oevelen, Chris; Graf, Thomas] Pompeu Fabra Univ, Dr Aiguader 88, Barcelona 08003, Spain; [Di Stefano, Bruno] Massachusetts Gen Hosp, Dept Mol Biol, Boston, MA 02114 USA; [Di Stefano, Bruno] Harvard Univ, Sch Med, Boston, MA 02114 USA; [de Wit, Elzo] Netherlands Canc Inst, Div Gene Regulat, Plesmanlaan 121, NL-1066 CX Amsterdam, Netherlands	de Laat, W (reprint author), Hubrecht Inst KNAW, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands.; de Laat, W (reprint author), Univ Med Ctr Utrecht, Uppsalalaan 8, NL-3584 CT Utrecht, Netherlands.; Graf, T (reprint author), Ctr Genom Regulat CRG, Gene Regulat Stem Cells & Canc Programme, Dr Aiguader 88, Barcelona 08003, Spain.; Graf, T (reprint author), Pompeu Fabra Univ, Dr Aiguader 88, Barcelona 08003, Spain.	w.delaat@hubrecht.eu; thomas.graf@crg.euw.delaat@hubrecht.eu; thomas.graf@crg.eu	Di Stefano, Bruno/K-5020-2017	Di Stefano, Bruno/0000-0003-2532-3087; Krijger, Peter/0000-0003-1702-348X; de Wit, Elzo/0000-0003-2883-1415; de Laat, Wouter/0000-0002-6393-595X	NWO/CW TOP grantNetherlands Organization for Scientific Research (NWO) [714.012.002]; NWO VICI grantNetherlands Organization for Scientific Research (NWO) [724.012.003]; NanoNextNL grant; European Research CouncilEuropean Research Council (ERC) [209700]; Ministerio de Educacion y CienciaSpanish Government [SAF.2012-37167]; Fundacio La Marato [TV3 120410]; AGAUR SGRAgencia de Gestio D'Ajuts Universitaris de Recerca Agaur (AGAUR) [1136]; European Research Council Synergy GrantEuropean Research Council (ERC); ERC Stg [637587]	This work was supported by an NWO/CW TOP grant (714.012.002), an NWO VICI grant 724.012.003, a NanoNextNL grant, and a European Research Council Starting Grant (209700, "4C") to W.d.L.; a Ministerio de Educacion y Ciencia, SAF.2012-37167, Fundacio La Marato TV3 120410, AGAUR SGR 1136, and European Research Council Synergy Grant ("4D-Genome) to T.G.; and an ERC Stg (637587, "HAP-PHEN") to E.d.W.	Apostolou E, 2013, CELL STEM CELL, V12, P699, DOI 10.1016/j.stem.2013.04.013; Bar-Nur O, 2011, CELL STEM CELL, V9, P17, DOI 10.1016/j.stem.2011.06.007; Boiani M, 2002, GENE DEV, V16, P1209, DOI 10.1101/gad.966002; Buganim Y, 2013, NAT REV GENET, V14, P427, DOI 10.1038/nrg3473; Buganim Y, 2012, CELL, V150, P1209, DOI 10.1016/j.cell.2012.08.023; Carey BW, 2010, NAT METHODS, V7, P56, DOI [10.1038/NMETH.1410, 10.1038/nmeth.1410]; Chandra T, 2015, CELL REP, V10, P471, DOI 10.1016/j.celrep.2014.12.055; Creyghton MP, 2010, P NATL ACAD SCI USA, V107, P21931, DOI 10.1073/pnas.1016071107; de Laat W, 2013, NATURE, V502, P499, DOI 10.1038/nature12753; de Wit E, 2013, NATURE, V501, P227, DOI 10.1038/nature12420; Denholtz M, 2013, CELL STEM CELL, V13, P602, DOI 10.1016/j.stem.2013.08.013; Di Stefano B, 2014, NATURE, V506, P235, DOI 10.1038/nature12885; Dixon JR, 2015, NATURE, V518, P331, DOI 10.1038/nature14222; Dixon JR, 2012, NATURE, V485, P376, DOI 10.1038/nature11082; Kagey MH, 2010, NATURE, V467, P430, DOI 10.1038/nature09380; Kim K, 2010, NATURE, V467, P285, DOI 10.1038/nature09342; Lane AA, 2014, NAT GENET, V46, P618, DOI 10.1038/ng.2949; Lieberman-Aiden E, 2009, SCIENCE, V326, P289, DOI 10.1126/science.1181369; Lodato MA, 2013, PLOS GENET, V9, DOI 10.1371/journal.pgen.1003288; Lujan E, 2015, NATURE, V521, P352, DOI 10.1038/nature14274; Nishino K, 2011, PLOS GENET, V7, DOI 10.1371/journal.pgen.1002085; Nora EP, 2012, NATURE, V485, P381, DOI 10.1038/nature11049; O'Malley J, 2013, NATURE, V499, P88, DOI 10.1038/nature12243; Peric-Hupkes D, 2010, MOL CELL, V38, P603, DOI 10.1016/j.molcel.2010.03.016; Phillips-Cremins JE, 2013, CELL, V153, P1281, DOI 10.1016/j.cell.2013.04.053; Polo JM, 2010, NAT BIOTECHNOL, V28, P848, DOI 10.1038/nbt.1667; Rao SSP, 2014, CELL, V159, P1665, DOI 10.1016/j.cell.2014.11.021; Sexton T, 2012, CELL, V148, P458, DOI 10.1016/j.cell.2012.01.010; Splinter E, 2012, METHODS, V58, P221, DOI 10.1016/j.ymeth.2012.04.009; Takahashi K, 2006, CELL, V126, P663, DOI 10.1016/j.cell.2006.07.024; van Oevelen C, 2008, MOL CELL, V32, P359, DOI 10.1016/j.molcel.2008.10.015; Wei Z, 2013, CELL STEM CELL, V13, P36, DOI 10.1016/j.stem.2013.05.010; Weissbein U, 2014, J CELL BIOL, V204, P153, DOI 10.1083/jcb.201310135; Whyte WA, 2013, CELL, V153, P307, DOI 10.1016/j.cell.2013.03.035; Yue F, 2014, NATURE, V515, P355, DOI 10.1038/nature13992; Zhang B, 2005, NUCLEIC ACIDS RES, V33, pW741, DOI 10.1093/nar/gki475; Zhao XY, 2009, NATURE, V461, P86, DOI 10.1038/nature08267; Zunder ER, 2015, CELL STEM CELL, V16, P323, DOI 10.1016/j.stem.2015.01.015	38	87	90	0	32	CELL PRESS	CAMBRIDGE	50 HAMPSHIRE ST, FLOOR 5, CAMBRIDGE, MA 02139 USA	1934-5909	1875-9777		CELL STEM CELL	Cell Stem Cell	MAY 5	2016	18	5					597	610		10.1016/j.stem.2016.01.007			14	Cell & Tissue Engineering; Cell Biology	Cell Biology	DL4HE	WOS:000375594200011	26971819	Other Gold, Green Published			2020-07-01	
J	Samusik, N; Good, Z; Spitzer, MH; Davis, KL; Nolan, GP				Samusik, Nikolay; Good, Zinaida; Spitzer, Matthew H.; Davis, Kara L.; Nolan, Garry P.			Automated mapping of phenotype space with single-cell data	NATURE METHODS			English	Article							FLOW-CYTOMETRY DATA; MASS CYTOMETRY; IDENTIFICATION; POPULATIONS; RESPONSES	Accurate identification of cell subsets in complex populations is key to discovering novelty in multidimensional single-cell experiments. We present X-shift (http://web.stanford.edu/similar to samusik/vortex/), an algorithm that processes data sets using fast k-nearest-neighbor estimation of cell event density and arranges populations by marker-based classification. X-shift enables automated cell-subset clustering and access to biological insights that 'prior knowledge' might prevent the researcher from discovering.	[Samusik, Nikolay; Good, Zinaida; Spitzer, Matthew H.; Davis, Kara L.; Nolan, Garry P.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Good, Zinaida; Spitzer, Matthew H.] Stanford Univ, Sch Med, Dept Pathol, Stanford, CA 94305 USA; [Davis, Kara L.] Stanford Univ, Sch Med, Dept Pediat Hematol & Oncol, Stanford, CA 94305 USA[Samusik, Nikolay; Good, Zinaida; Spitzer, Matthew H.; Davis, Kara L.; Nolan, Garry P.] Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA; [Good, Zinaida; Spitzer, Matthew H.] Stanford Univ, Sch Med, Dept Pathol, Stanford, CA 94305 USA; [Davis, Kara L.] Stanford Univ, Sch Med, Dept Pediat Hematol & Oncol, Stanford, CA 94305 USA	Nolan, GP (reprint author), Stanford Univ, Sch Med, Dept Microbiol & Immunol, Stanford, CA 94305 USA.	gnolan@stanford.edugnolan@stanford.edu	Nolan, Garry/AAE-7903-2019	Spitzer, Matthew/0000-0002-5291-3819; Good, Zinaida/0000-0003-2343-5771	NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [R01GM109836, U19 AI057229, 1U19AI100627, R01-CA184968, 1R33-CA183654-01, R33-CA183692, 1R01-GM10983601, 1R21-CA183660, 1R01-NS08953301, OPP-1113682, 5UH2-AR067676, 1R01-CA19665701, R01-HL120724]; Immunology Training grant [5T32AI007290]; US Department of DefenseUnited States Department of Defense; Northrop-Grumman Corporation; US Food and Drug Administration [BAA HHSF223201210194c]; Rachford and Carlota A. Harris Endowed Chair	We thank M. Angst, W.J. Fantl, A. Surnov, E. Freeman, and W.H. Wong for help in manuscript editing and preparation. This work was supported by NIH grant R01GM109836 (N.S.); NIH grants U19 AI057229, 1U19AI100627, R01-CA184968, 1R33-CA183654-01, R33-CA183692, 1R01-GM10983601, 1R21-CA183660, 1R01-NS08953301, OPP-1113682, 5UH2-AR067676, 1R01-CA19665701 and R01-HL120724 (G.P.N.); Immunology Training grant 5T32AI007290 (Z.G.); US Department of Defense Teal Innovator Award (G.P.N.); Northrop-Grumman Corporation (G.P.N.); the US Food and Drug Administration grant BAA HHSF223201210194c (G.P.N.) and the Rachford and Carlota A. Harris Endowed Chair (G.P.N.).	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Methods	JUN	2016	13	6					493	+		10.1038/NMETH.3863			7	Biochemical Research Methods	Biochemistry & Molecular Biology	DO0PF	WOS:000377480100015	27183440	Green Accepted			2020-07-01	
J	Prill, RJ; Saez-Rodriguez, J; Alexopoulos, LG; Sorger, PK; Stolovitzky, G				Prill, Robert J.; Saez-Rodriguez, Julio; Alexopoulos, Leonidas G.; Sorger, Peter K.; Stolovitzky, Gustavo			Crowdsourcing Network Inference: The DREAM Predictive Signaling Network Challenge	SCIENCE SIGNALING			English	Article							MASS CYTOMETRY; GENE NETWORKS; CELL; RECEPTOR	Computational analyses of systematic measurements on the states and activities of signaling proteins (as captured by phosphoproteomic data, for example) have the potential to uncover uncharacterized protein-protein interactions and to identify the subset that are important for cellular response to specific biological stimuli. However, inferring mechanistically plausible protein signaling networks (PSNs) from phosphoproteomics data is a difficult task, owing in part to the lack of sufficiently comprehensive experimental measurements, the inherent limitations of network inference algorithms, and a lack of standards for assessing the accuracy of inferred PSNs. A case study in which 12 research groups inferred PSNs from a phosphoproteomics data set demonstrates an assessment of inferred PSNs on the basis of the accuracy of their predictions. The concurrent prediction of the same previously unreported signaling interactions by different participating teams suggests relevant validation experiments and establishes a framework for combining PSNs inferred by multiple research groups into a composite PSN. We conclude that crowdsourcing the construction of PSNs-that is, outsourcing the task to the interested community-may be an effective strategy for network inference.	[Prill, Robert J.; Stolovitzky, Gustavo] IBM Computat Biol Ctr, Yorktown Hts, NY 10598 USA; [Prill, Robert J.; Saez-Rodriguez, Julio] European Bioinformat Inst EMBL EBI, Cambridge CB10 1SD, England; [Saez-Rodriguez, Julio] European Mol Biol Lab, Genome Biol Unit, D-69117 Heidelberg, Germany; [Alexopoulos, Leonidas G.] Natl Tech Univ Athens, Dept Mech Engn, Athens 15780, Greece; [Sorger, Peter K.] Harvard Univ, Sch Med, Dept Syst Biol, Boston, MA 02115 USA; [Sorger, Peter K.] MIT, Dept Biol Engn, Cambridge, MA 02139 USA[Prill, Robert J.; Stolovitzky, Gustavo] IBM Computat Biol Ctr, Yorktown Hts, NY 10598 USA; [Prill, Robert J.; Saez-Rodriguez, Julio] European Bioinformat Inst EMBL EBI, Cambridge CB10 1SD, England; [Saez-Rodriguez, Julio] European Mol Biol Lab, Genome Biol Unit, D-69117 Heidelberg, Germany; [Alexopoulos, Leonidas G.] Natl Tech Univ Athens, Dept Mech Engn, Athens 15780, Greece; [Sorger, Peter K.] Harvard Univ, Sch Med, Dept Syst Biol, Boston, MA 02115 USA; [Sorger, Peter K.] MIT, Dept Biol Engn, Cambridge, MA 02139 USA	Stolovitzky, G (reprint author), IBM Computat Biol Ctr, Yorktown Hts, NY 10598 USA.	gustavo@us.ibm.comgustavo@us.ibm.com	Saez-Rodriguez, Julio/H-7114-2019	Saez-Rodriguez, Julio/0000-0002-8552-8976; Sorger, Peter/0000-0002-3364-1838	National Institutes of Health (NIH) through the Columbia University Center for Multiscale Analysis Genomic and Cellular Networks (MAGNet)United States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Neurological Disorders & Stroke (NINDS); LINCS [U54HG006097];  [GM68762]	We acknowledge support for the DREAM project by the National Institutes of Health (NIH) Roadmap Initiative, through the Columbia University Center for Multiscale Analysis Genomic and Cellular Networks (MAGNet), and grant GM68762 for data generation; the LINCS program is currently funding data collection for future DREAM challenges (U54HG006097).	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Signal.	SEP 6	2011	4	189							mr7	10.1126/scisignal.2002212			6	Biochemistry & Molecular Biology; Cell Biology	Biochemistry & Molecular Biology; Cell Biology	816LQ	WOS:000294601900003	21900204	Green Accepted			2020-07-01	
J	Olin, A; Henckel, E; Chen, Y; Lakshmikanth, T; Pou, C; Mikes, J; Gustafsson, A; Bernhardsson, AK; Zhang, C; Bohlin, K; Brodin, P				Olin, Axel; Henckel, Ewa; Chen, Yang; Lakshmikanth, Tadepally; Pou, Christian; Mikes, Jaromir; Gustafsson, Anna; Bernhardsson, Anna Karin; Zhang, Cheng; Bohlin, Kajsa; Brodin, Petter			Stereotypic Immune System Development in Newborn Children	CELL			English	Article							YELLOW-FEVER VACCINE; MASS CYTOMETRY; T-CELLS; BACTERIAL-INFECTIONS; SUPPRESSOR-CELLS; COLONIZATION; POPULATIONS; MICROBIOTA; RESPONSES; BIOLOGY	Epidemiological data suggest that early life exposures are key determinants of immune-mediated disease later in life. Young children are also particularly susceptible to infections, warranting more analyses of immune system development early in life. Such analyses mostly have been performed in mouse models or human cord blood samples, but these cannot account for the complex environmental exposures influencing human newborns after birth. Here, we performed longitudinal analyses in 100 newborn children, sampled up to 4 times during their first 3 months of life. From 100 mu L of blood, we analyze the development of 58 immune cell populations by mass cytometry and 267 plasma proteins by immunoassays, uncovering drastic changes not predictable from cord blood measurements but following a stereotypic pattern. Preterm and term children differ at birth but converge onto a shared trajectory, seemingly driven by microbial interactions and hampered by early gut bacterial dysbiosis.	[Olin, Axel; Chen, Yang; Lakshmikanth, Tadepally; Pou, Christian; Mikes, Jaromir; Bernhardsson, Anna Karin; Brodin, Petter] Karolinska Inst, Dept Womens & Childrens Hlth, Sci Life Lab, S-17121 Solna, Sweden; [Henckel, Ewa; Gustafsson, Anna; Bohlin, Kajsa] Karolinska Inst, Dept Clin Sci Intervent & Technol, S-14152 Solna, Sweden; [Henckel, Ewa; Gustafsson, Anna; Bernhardsson, Anna Karin; Bohlin, Kajsa; Brodin, Petter] Karolinska Univ Hosp, Dept Neonatol, S-17176 Solna, Sweden; [Zhang, Cheng] KTH, Royal Inst Technol, Sch Biotechnol, Sci Life Lab, S-17121 Stockholm, Sweden[Olin, Axel; Chen, Yang; Lakshmikanth, Tadepally; Pou, Christian; Mikes, Jaromir; Bernhardsson, Anna Karin; Brodin, Petter] Karolinska Inst, Dept Womens & Childrens Hlth, Sci Life Lab, S-17121 Solna, Sweden; [Henckel, Ewa; Gustafsson, Anna; Bohlin, Kajsa] Karolinska Inst, Dept Clin Sci Intervent & Technol, S-14152 Solna, Sweden; [Henckel, Ewa; Gustafsson, Anna; Bernhardsson, Anna Karin; Bohlin, Kajsa; Brodin, Petter] Karolinska Univ Hosp, Dept Neonatol, S-17176 Solna, Sweden; [Zhang, Cheng] KTH, Royal Inst Technol, Sch Biotechnol, Sci Life Lab, S-17121 Stockholm, Sweden	Brodin, P (reprint author), Karolinska Inst, Dept Womens & Childrens Hlth, Sci Life Lab, S-17121 Solna, Sweden.; Brodin, P (reprint author), Karolinska Univ Hosp, Dept Neonatol, S-17176 Solna, Sweden.	petter.brodin@ki.sepetter.brodin@ki.se	Brodin, Petter/C-3317-2011; Mikes, Jaromir/X-3588-2019; Henckel, Ewa/AAD-3185-2019; Zhang, Cheng/L-7906-2016; Tadepally, Lakshmikanth/P-5154-2019	Brodin, Petter/0000-0002-8103-0046; Mikes, Jaromir/0000-0002-9941-7855; Zhang, Cheng/0000-0002-3721-8586; Tadepally, Lakshmikanth/0000-0001-7256-5770; Olin, Axel/0000-0002-1161-4476; Henckel, Ewa/0000-0003-0775-1563	ERCEuropean Research Council (ERC) [StG-677559]; Swedish Research Council,Swedish Research Council; Karolinska InstitutetKarolinska Institutet; Swedish Society for Medical Research	We thank all families for their participation and nurses, midwives, and physicians for help with sample collection. We thank Jochen Schwenk and Ragna ussler at the SciLifeLab Plasma profiling platform and Lars Engstrand and Luisa Hugerth at the Clinical Genomics Facility for help with 16S rRNA profiling. P.B. is grateful to Professor Mark Davis of Stanford University for his valuable mentorship. This study was supported by an ERC Starting Grant (StG-677559), the Swedish Research Council, Karolinska Institutet, and the Swedish Society for Medical Research.	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J	Chen, H; Lau, MC; Wong, MT; Newell, EW; Poidinger, M; Chen, JM				Chen, Hao; Lau, Mai Chan; Wong, Michael Thomas; Newell, Evan W.; Poidinger, Michael; Chen, Jinmiao			Cytofkit: A Bioconductor Package for an Integrated Mass Cytometry Data Analysis Pipeline	PLOS COMPUTATIONAL BIOLOGY			English	Article							CELL; PROGRESSION	Single-cell mass cytometry significantly increases the dimensionality of cytometry analysis as compared to fluorescence flow cytometry, providing unprecedented resolution of cellular diversity in tissues. However, analysis and interpretation of these high-dimensional data poses a significant technical challenge. Here, we present cytofkit, a new Bioconductor package, which integrates both state-of-the-art bioinformatics methods and in-house novel algorithms to offer a comprehensive toolset for mass cytometry data analysis. Cytofkit provides functions for data pre-processing, data visualization through linear or non-linear dimensionality reduction, automatic identification of cell subsets, and inference of the relatedness between cell subsets. This pipeline also provides a graphical user interface (GUI) for ease of use, as well as a shiny application (APP) for interactive visualization of cell subpopulations and progression profiles of key markers. Applied to a CD14(-)CD19(-) PBMCs dataset, cytofkit accurately identified different subsets of lymphocytes; applied to a human CD4(+) T cell dataset, cytofkit uncovered multiple subtypes of TFH cells spanning blood and tonsils. Cytofkit is implemented in R, licensed under the Artistic license 2.0, and freely available from the Bioconductor website, https://bioconductor.org/packages/cytofkit/. Cytofkit is also applicable for flow cytometry data analysis.	[Chen, Hao; Lau, Mai Chan; Wong, Michael Thomas; Newell, Evan W.; Poidinger, Michael; Chen, Jinmiao] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore[Chen, Hao; Lau, Mai Chan; Wong, Michael Thomas; Newell, Evan W.; Poidinger, Michael; Chen, Jinmiao] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore	Chen, JM (reprint author), ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore.	chen_jinmiao@immunol.a-star.edu.sgchen_jinmiao@immunol.a-star.edu.sg	Newell, Evan/AAE-9470-2020; Newell, Evan W/F-9711-2012; Lau, Mai Chan/L-5186-2019	Newell, Evan/0000-0002-2889-243X; Newell, Evan W/0000-0002-2889-243X; chen, jinmiao/0000-0001-7547-6423; Poidinger, Michael/0000-0002-1047-2277	A-STAR/SIgNAgency for Science Technology & Research (ASTAR)	This study was funded by A-STAR/SIgN core funding (JC). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.	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Biol.	SEP	2016	12	9							e1005112	10.1371/journal.pcbi.1005112			17	Biochemical Research Methods; Mathematical & Computational Biology	Biochemistry & Molecular Biology; Mathematical & Computational Biology	EB8RN	WOS:000387658600032	27662185	DOAJ Gold, Green Published			2020-07-01	
J	Newell, EW; Davis, MM				Newell, Evan W.; Davis, Mark M.			Beyond model antigens: high-dimensional methods for the analysis of antigen-specific T cells	NATURE BIOTECHNOLOGY			English	Review							MASS CYTOMETRY; COMPREHENSIVE ANALYSIS; PARALLEL DETECTION; GENE-EXPRESSION; FLOW-CYTOMETRY; RESPONSES; VIRUS; MEMORY; IMMUNE; REPERTOIRE	Adaptive immune responses often begin with the formation of a molecular complex between a T-cell receptor (TCR) and a peptide antigen bound to a major histocompatibility complex (MHC) molecule. These complexes are highly variable, however, due to the polymorphism of MHC genes, the random, inexact recombination of TCR gene segments, and the vast array of possible self and pathogen peptide antigens. As a result, it has been very difficult to comprehensively study the TCR repertoire or identify and track more than a few antigen-specific T cells in mice or humans. For mouse studies, this had led to a reliance on model antigens and TCR transgenes. The study of limited human clinical samples, in contrast, requires techniques that can simultaneously survey TCR phenotype and function, and TCR reactivity to many T-cell epitopes. Thanks to recent advances in single-cell and cytometry methodologies, as well as high-throughput sequencing of the TCR repertoire, we now have or will soon have the tools needed to comprehensively analyze T-cell responses in health and disease.	[Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Davis, Mark M.] Howard Hughes Med Inst, Dept Microbiol & Immunol, Stanford, CA USA; [Davis, Mark M.] Howard Hughes Med Inst, Inst Immun Transplantat & Infect, Stanford, CA USA; [Davis, Mark M.] Howard Hughes Med Inst, Stanford, CA USA[Newell, Evan W.] ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore; [Davis, Mark M.] Howard Hughes Med Inst, Dept Microbiol & Immunol, Stanford, CA USA; [Davis, Mark M.] Howard Hughes Med Inst, Inst Immun Transplantat & Infect, Stanford, CA USA; [Davis, Mark M.] Howard Hughes Med Inst, Stanford, CA USA	Newell, EW (reprint author), ASTAR, Singapore Immunol Network SIgN, Singapore, Singapore.	evan_newell@immunol.a-star.edu.sg; mmdavis@stanford.eduevan_newell@immunol.a-star.edu.sg; mmdavis@stanford.edu	Newell, Evan/AAE-9470-2020; Newell, Evan W/F-9711-2012	Newell, Evan/0000-0002-2889-243X; Newell, Evan W/0000-0002-2889-243X	Howard Hughes Medical InstituteHoward Hughes Medical Institute; NIAID NIH HHSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Allergy & Infectious Diseases (NIAID) [U19 AI057229, R01 AI022511]		Adams JJ, 2011, IMMUNITY, V35, P681, DOI 10.1016/j.immuni.2011.09.013; Aghaeepour N, 2013, NAT METHODS, V10, P228, DOI [10.1038/NMETH.2365, 10.1038/nmeth.2365]; Akram A, 2012, CLIN IMMUNOL, V143, P99, DOI 10.1016/j.clim.2012.01.015; Altman JD, 1996, SCIENCE, V274, P94, DOI 10.1126/science.274.5284.94; Amir ED, 2013, NAT BIOTECHNOL, V31, P545, DOI 10.1038/nbt.2594; Andersen RS, 2012, NAT PROTOC, V7, P891, DOI 10.1038/nprot.2012.037; Arstila TP, 1999, SCIENCE, V286, P958, DOI 10.1126/science.286.5441.958; Assarsson E, 2008, J VIROL, V82, P12241, DOI 10.1128/JVI.01563-08; Baker ES, 2012, GENOME MED, V4, DOI 10.1186/gm364; Bendall SC, 2011, SCIENCE, V332, P687, DOI 10.1126/science.1198704; Betts MR, 2006, BLOOD, V107, P4781, DOI 10.1182/blood-2005-12-4818; Betts MR, 2003, J IMMUNOL METHODS, V281, P65, DOI 10.1016/S0022-1759(03)00265-5; Birnbaum ME, 2012, IMMUNOL REV, V250, P82, DOI 10.1111/imr.12006; Bjornson ZB, 2013, CURR OPIN IMMUNOL, V25, P484, DOI 10.1016/j.coi.2013.07.004; Bodenmiller B, 2012, NAT BIOTECHNOL, V30, P858, DOI 10.1038/nbt.2317; Bowen DG, 2005, NATURE, V436, P946, DOI 10.1038/nature04079; Boyd SD, 2013, CURR OPIN IMMUNOL, V25, P511, DOI 10.1016/j.coi.2013.07.007; BRUNNER KT, 1968, IMMUNOLOGY, V14, P181; BURNET F. 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Biotechnol.	FEB	2014	32	2					149	157		10.1038/nbt.2783			9	Biotechnology & Applied Microbiology	Biotechnology & Applied Microbiology	AA4NY	WOS:000331074000020	24441473	Green Accepted			2020-07-01	
J	Strauss-Albee, DM; Fukuyama, J; Liang, EC; Yao, Y; Jarrell, JA; Drake, AL; Kinuthia, J; Montgomery, RR; John-Stewart, G; Holmes, S; Blish, CA				Strauss-Albee, Dara M.; Fukuyama, Julia; Liang, Emily C.; Yao, Yi; Jarrell, Justin A.; Drake, Alison L.; Kinuthia, John; Montgomery, Ruth R.; John-Stewart, Grace; Holmes, Susan; Blish, Catherine A.			Human NK cell repertoire diversity reflects immune experience and correlates with viral susceptibility	SCIENCE TRANSLATIONAL MEDICINE			English	Article							NATURAL-KILLER-CELLS; NILE VIRUS-INFECTION; MASS CYTOMETRY; HIV ACQUISITION; CUTTING EDGE; T-CELLS; EXPRESSION; RESPONSES; INNATE; CYTOMEGALOVIRUS	Innate natural killer (NK) cells are diverse at the single-cell level because of variegated expressions of activating and inhibitory receptors, yet the developmental roots and functional consequences of this diversity remain unknown. Because NK cells are critical for antiviral and antitumor responses, a better understanding of their diversity could lead to an improved ability to harness them therapeutically. We found that NK diversity is lower at birth than in adults. During an antiviral response to either HIV-1 or West Nile virus, NK diversity increases, resulting in terminal differentiation and cytokine production at the cost of cell division and degranulation. In African women matched for HIV-1 exposure risk, high NK diversity is associated with increased risk of HIV-1 acquisition. Existing diversity may therefore decrease the flexibility of the antiviral response. Collectively, the data reveal that human NK diversity is a previously undefined metric of immune history and function that may be clinically useful in forecasting the outcomes of infection and malignancy.	[Strauss-Albee, Dara M.; Jarrell, Justin A.; Blish, Catherine A.] Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA; [Strauss-Albee, Dara M.; Liang, Emily C.; Jarrell, Justin A.; Blish, Catherine A.] Stanford Univ, Sch Med, Dept Med, Stanford, CA 94305 USA; [Fukuyama, Julia; Holmes, Susan] Stanford Univ, Dept Stat, Stanford, CA 94305 USA; [Yao, Yi; Montgomery, Ruth R.] Yale Univ, Sch Med, Dept Internal Med, New Haven, CT 06519 USA; [Drake, Alison L.; John-Stewart, Grace] Univ Washington, Dept Global Hlth, Seattle, WA 98195 USA; [Kinuthia, John] Kenyatta Natl Hosp, Dept Res & Programs, Nairobi 00202, Kenya; [John-Stewart, Grace] Univ Washington, Dept Epidemiol, Seattle, WA 98195 USA; [John-Stewart, Grace] Univ Washington, Dept Med, Seattle, WA 98195 USA; [John-Stewart, Grace] Univ Washington, Dept Pediat, Seattle, WA 98195 USA[Strauss-Albee, Dara M.; Jarrell, Justin A.; Blish, Catherine A.] Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA; [Strauss-Albee, Dara M.; Liang, Emily C.; Jarrell, Justin A.; Blish, Catherine A.] Stanford Univ, Sch Med, Dept Med, Stanford, CA 94305 USA; [Fukuyama, Julia; Holmes, Susan] Stanford Univ, Dept Stat, Stanford, CA 94305 USA; [Yao, Yi; Montgomery, Ruth R.] Yale Univ, Sch Med, Dept Internal Med, New Haven, CT 06519 USA; [Drake, Alison L.; John-Stewart, Grace] Univ Washington, Dept Global Hlth, Seattle, WA 98195 USA; [Kinuthia, John] Kenyatta Natl Hosp, Dept Res & Programs, Nairobi 00202, Kenya; [John-Stewart, Grace] Univ Washington, Dept Epidemiol, Seattle, WA 98195 USA; [John-Stewart, Grace] Univ Washington, Dept Med, Seattle, WA 98195 USA; [John-Stewart, Grace] Univ Washington, Dept Pediat, Seattle, WA 98195 USA	Blish, CA (reprint author), Stanford Univ, Sch Med, Stanford Immunol, Stanford, CA 94305 USA.	cblish@stanford.educblish@stanford.edu		Blish, Catherine/0000-0001-6946-7627; John-Stewart, Grace/0000-0002-4301-1573; Holmes, Susan/0000-0002-2208-8168; Montgomery, Ruth/0000-0002-8661-4454	National Science FoundationNational Science Foundation (NSF) [DGE-114740]; Ruth L. Kirschstein National Research Service AwardUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [1F31AI118469-01]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [U19AI089992, DP2AI11219301, P01 HSD 064915]; Beckman Young Investigator Award; California HIV Research Program IDEA award; University of Washington Center for AIDS Research [P30 AI27757]	Funding for this work was provided by National Science Foundation training grant DGE-114740 (D.M.S.-A.), Ruth L. Kirschstein National Research Service Award 1F31AI118469-01 (D.M.S.-A.), NIH Award U19AI089992 (R.R.M.), a Beckman Young Investigator Award (C.A.B.), a California HIV Research Program IDEA award (C.A.B.), and NIH Directors' New Innovator Award DP2AI11219301 (C.A.B.). The Kenyan studies were funded by NIH (P01 HSD 064915) and received assistance from the University of Washington Center for AIDS Research (P30 AI27757).	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Transl. Med.	JUL 22	2015	7	297							297ra115	10.1126/scitranslmed.aac5722			11	Cell Biology; Medicine, Research & Experimental	Cell Biology; Research & Experimental Medicine	CN9AT	WOS:000358738700007	26203083	Green Accepted, Bronze			2020-07-01	
J	Saadatpour, A; Lai, SJ; Guo, GJ; Yuan, GC				Saadatpour, Assieh; Lai, Shujing; Guo, Guoji; Yuan, Guo-Cheng			Single-Cell Analysis in Cancer Genomics	TRENDS IN GENETICS			English	Review							GENE-EXPRESSION ANALYSIS; CIRCULATING TUMOR-CELLS; EMBRYONIC STEM-CELLS; MESSENGER-RNA-SEQ; MASS CYTOMETRY; TRANSCRIPTOME ANALYSIS; NETWORK ANALYSIS; FATE DECISIONS; IN-SITU; T-SNE	Genetic changes and environmental differences result in cellular heterogeneity among cancer cells within the same tumor, thereby complicating treatment outcomes. Recent advances in single-cell technologies have opened new avenues to characterize the intra-tumor cellular heterogeneity, identify rare cell types, measure mutation rates, and, ultimately, guide diagnosis and treatment. In this paper we review the recent single-cell technological and computational advances at the genomic, transcriptomic, and proteomic levels, and discuss their applications in cancer research.	[Saadatpour, Assieh; Yuan, Guo-Cheng] Dana Farber Canc Inst, Dept Biostat & Computat Biol, Boston, MA 02215 USA; [Saadatpour, Assieh; Yuan, Guo-Cheng] Harvard Univ, TH Chan Sch Publ Hlth, Dept Biostat, Boston, MA 02115 USA; [Lai, Shujing; Guo, Guoji] Zhejiang Univ, Sch Med, Ctr Stem Cell & Regenerat Med, Hangzhou 310058, Zhejiang, Peoples R China; [Yuan, Guo-Cheng] Harvard Stem Cell Inst, Cambridge, MA 02138 USA[Saadatpour, Assieh; Yuan, Guo-Cheng] Dana Farber Canc Inst, Dept Biostat & Computat Biol, Boston, MA 02215 USA; [Saadatpour, Assieh; Yuan, Guo-Cheng] Harvard Univ, TH Chan Sch Publ Hlth, Dept Biostat, Boston, MA 02115 USA; [Lai, Shujing; Guo, Guoji] Zhejiang Univ, Sch Med, Ctr Stem Cell & Regenerat Med, Hangzhou 310058, Zhejiang, Peoples R China; [Yuan, Guo-Cheng] Harvard Stem Cell Inst, Cambridge, MA 02138 USA	Guo, GJ (reprint author), Zhejiang Univ, Sch Med, Ctr Stem Cell & Regenerat Med, Hangzhou 310058, Zhejiang, Peoples R China.	ggj@zju.edu.cn; gcyuan@jimmy.harvard.eduggj@zju.edu.cn; gcyuan@jimmy.harvard.edu	t, lf/AAJ-4924-2020		Claudia Adams Barr Award	This work was supported in part by a Claudia Adams Barr Award to G-C.Y.	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OCT	2015	31	10			SI		576	586		10.1016/j.tig.2015.07.003			11	Genetics & Heredity	Genetics & Heredity	CU2JN	WOS:000363349900005	26450340	Green Accepted			2020-07-01	
J	Diggins, KE; Ferrell, PB; Irish, JM				Diggins, Kirsten E.; Ferrell, P. Brent, Jr.; Irish, Jonathan M.			Methods for discovery and characterization of cell subsets in high dimensional mass cytometry data	METHODS			English	Article						Mass cytometry; Flow cytometry; Single cell biology; Unsupervised analysis; Machine learning	FLOW-CYTOMETRY; AUTOMATED IDENTIFICATION; CLASSIFICATION; EXPRESSION; REDUCTION; NETWORKS; IMMUNE	The flood of high-dimensional data resulting from mass cytometry experiments that measure more than 40 features of individual cells has stimulated creation of new single cell computational biology tools. These tools draw on advances in the field of machine learning to capture multi-parametric relationships and reveal cells that are easily overlooked in traditional analysis. Here, we introduce a workflow for high dimensional mass cytometry data that emphasizes unsupervised approaches and visualizes data in both single cell and population level views. This workflow includes three central components that are common across mass cytometty analysis approaches: (1) distinguishing initial populations, (2) revealing cell subsets, and (3) characterizing subset features. In the implementation described here, viSNE, SPADE, and heatmaps were used sequentially to comprehensively characterize and compare healthy and malignant human tissue samples. The use of multiple methods helps provide a comprehensive view of results, and the largely unsupervised workflow facilitates automation and helps researchers avoid missing cell populations with unusual or unexpected phenotypes. Together, these methods develop a framework for future machine learning of cell identity. (C) 2015 Elsevier Inc. All rights reserved.	[Diggins, Kirsten E.; Irish, Jonathan M.] Vanderbilt Univ, Sch Med, Canc Biol, Nashville, TN 37232 USA; [Ferrell, P. Brent, Jr.] Vanderbilt Univ, Sch Med, Med Div Hematol Oncol, Nashville, TN 37232 USA; [Irish, Jonathan M.] Vanderbilt Univ, Sch Med, Pathol Microbiol & Immunol, Nashville, TN 37232 USA[Diggins, Kirsten E.; Irish, Jonathan M.] Vanderbilt Univ, Sch Med, Canc Biol, Nashville, TN 37232 USA; [Ferrell, P. Brent, Jr.] Vanderbilt Univ, Sch Med, Med Div Hematol Oncol, Nashville, TN 37232 USA; [Irish, Jonathan M.] Vanderbilt Univ, Sch Med, Pathol Microbiol & Immunol, Nashville, TN 37232 USA	Irish, JM (reprint author), Vanderbilt Univ, Sch Med, 740B Preston Bldg,2220 Pierce Ave, Nashville, TN 37232 USA.	jonathan.irish@vanderbilt.edujonathan.irish@vanderbilt.edu	Irish, Jonathan M/E-1180-2013; Diggins, Kirsten/F-7348-2015	Irish, Jonathan M/0000-0001-9428-8866; Ferrell, Paul/0000-0003-1140-9154; Diggins, Kirsten/0000-0003-1622-0158	Vanderbilt-Ingram Cancer Center (VICC) [P30 CA68485]; VICC Young Ambassadors award; VICC Hematology Helping Hands award; NIH/NCIUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [K12 CA090625];  [R25 CA136440-04]	This study was supported by R25 CA136440-04 (K.E.D.), NIH/NCI K12 CA090625 (P.B.F.), R00 CA143231-03 (J.M.I.), the Vanderbilt-Ingram Cancer Center (VICC, P30 CA68485), and VICC Young Ambassadors and VICC Hematology Helping Hands awards. 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L., 2015, CYTOMETRY A; van der Maaten L, 2008, J MACH LEARN RES, V9, P2579; Van Gassen S., 2015, CYTOMETRY A; Zare H, 2010, BMC BIOINFORMATICS, V11, DOI 10.1186/1471-2105-11-403	62	72	72	0	28	ACADEMIC PRESS INC ELSEVIER SCIENCE	SAN DIEGO	525 B ST, STE 1900, SAN DIEGO, CA 92101-4495 USA	1046-2023	1095-9130		METHODS	Methods	JUL 1	2015	82						55	63		10.1016/j.ymeth.2015.05.008			9	Biochemical Research Methods; Biochemistry & Molecular Biology	Biochemistry & Molecular Biology	CL4GN	WOS:000356910700008	25979346	Green Accepted			2020-07-01	
J	Miner, JJ; Yeang, HXA; Fox, JM; Taffner, S; Malkova, ON; Oh, ST; Kim, AHJ; Diamond, MS; Lenschow, DJ; Yokoyama, WM				Miner, Jonathan J.; Yeang, Han Xian Aw; Fox, Julie M.; Taffner, Samantha; Malkova, Olga N.; Oh, Stephen T.; Kim, Alfred H. J.; Diamond, Michael S.; Lenschow, Deborah J.; Yokoyama, Wayne M.			Chikungunya Viral Arthritis in the United States A Mimic of Seronegative Rheumatoid Arthritis	ARTHRITIS & RHEUMATOLOGY			English	Article							WESTERN-HEMISPHERE; VIRUS; DISEASE; SPREAD	Objective. Chikungunya virus (CHIKV) is an arthritogenic mosquito-transmitted alphavirus that spread to the Caribbean in 2013 and to the US in 2014. CHIKV-infected patients develop inflammatory arthritis that can persist for months or years, but little is known about the rheumatologic and immunologic features of CHIKV-related arthritis in humans, particularly as compared to rheumatoid arthritis (RA). The purpose of this study was to describe these features in a group of 10 American travelers who were nearly simultaneously infected while visiting Haiti in June 2014. Methods. Patient history was obtained and physical examination and laboratory tests were performed. All patients with CHIKV-related arthritis had detectable levels of anti-CHIKV IgG. Using cytometry by time-of-flight (CyTOF), we analyzed peripheral blood mononuclear cells in CHIKV-infected patients, healthy controls, and patients with untreated, active RA. Results. Among 10 CHIKV-infected individuals, 8 developed persistent symmetric polyarthritis that met the American College of Rheumatology/European League Against Rheumatism 2010 criteria for (seronegative) RA. CyTOF analysis revealed that RA and CHIKV-infected patients had greater percentages of activated and effector CD4+ and CD8+ T cells than healthy controls. Conclusion. In addition to similar clinical features, patients with CHIKV infection and patients with RA develop very similar peripheral T cell phenotypes. These overlapping clinical and immunologic features highlight a need for rheumatologists to consider CHIKV infection when evaluating patients with new, symmetric polyarthritis.	[Miner, Jonathan J.; Yeang, Han Xian Aw; Fox, Julie M.; Taffner, Samantha; Malkova, Olga N.; Oh, Stephen T.; Kim, Alfred H. J.; Diamond, Michael S.; Lenschow, Deborah J.; Yokoyama, Wayne M.] Washington Univ, Sch Med, St Louis, MO 63110 USA; [Yokoyama, Wayne M.] Howard Hughes Med Inst, Chevy Chase, MD USA[Miner, Jonathan J.; Yeang, Han Xian Aw; Fox, Julie M.; Taffner, Samantha; Malkova, Olga N.; Oh, Stephen T.; Kim, Alfred H. J.; Diamond, Michael S.; Lenschow, Deborah J.; Yokoyama, Wayne M.] Washington Univ, Sch Med, St Louis, MO 63110 USA; [Yokoyama, Wayne M.] Howard Hughes Med Inst, Chevy Chase, MD USA	Yokoyama, WM (reprint author), Washington Univ, Sch Med, Div Rheumatol, Campus Box 8045, St Louis, MO 63110 USA.	yokoyama@wustl.eduyokoyama@wustl.edu	Diamond, Michael/AAH-1733-2019; Kim, Alfred H.J./F-5699-2013	Diamond, Michael/0000-0002-8791-3165; Kim, Alfred H.J./0000-0003-4074-0516; Fox, Julie/0000-0003-0567-738X	Barnes-Jewish Hospital Foundation; Howard Hughes Medical InstituteHoward Hughes Medical Institute; Center for Human Immunology and Immunotherapy Programs at Washington University; NIH (Rhematic Diseases Core Center) [P30-AR-048335]; PfizerPfizer; JanssenJohnson & Johnson USAJanssen Biotech Inc; AmgenAmgen; InBios; MerckMerck & Company	Supported by the Barnes-Jewish Hospital Foundation, the Howard Hughes Medical Institute, the Center for Human Immunology and Immunotherapy Programs at Washington University, and the NIH (Rhematic Diseases Core Center grant P30-AR-048335).; Dr. Kim has received consulting fees, speaking fees, and/or honoraria from Pfizer, Janssen, and Amgen (less than $10,000 each). Dr. Diamond has received consulting fees from InBios and Merck (less than $10,000 each) and licensing fees and royalties from Merck for the use of monoclonal antibodies. Dr. Lenschow has received royalties from eBioscience and Santa Cruz Biotechnology for antibodies unrelated to this study.	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MAY	2015	67	5					1214	1220		10.1002/art.39027			7	Rheumatology	Rheumatology	CH1IV	WOS:000353775900011	25605621	Green Accepted			2020-07-01