Isotropic Imaging Across Spatial Scales with Axially Swept Light-Sheet Microscopy
Creators
- 1. Lyda Hill Department of Bioinformatics, University of Texas Southwestern Medical Center.
- 2. Department of Physics and Astronomy, University of New Mexico.
- 3. Coleman Technologies, Inc. D.B.A., Sciotex.
- 4. Department of Cell Biology, Yale University.
- 5. Department of Molecular Life Sciences, University of Zurich.
Description
Light-Sheet Fluorescence Microscopy (LSFM) is a rapidly growing technique that has gained tremendous popularity in the life sciences owing to its high-spatiotemporal resolution and gentle, non-phototoxic illumination. In this protocol, we provide detailed directions for the assembly and operation of a versatile LSFM variant, referred to as Axially Swept Light-Sheet Microscopy (ASLM), that delivers an unparalleled combination of field of view, optical resolution, and optical sectioning. To democratize ASLM, we provide an overview of its working principle, applications to biological imaging, as well as pragmatic tips for the assembly, alignment, and control of its optical systems. Furthermore, we provide detailed part lists and schematics for several variants of ASLM, that together, can resolve molecular detail in chemically expanded samples, subcellular organization in living cells, or the anatomical composition of chemically cleared intact organisms. We also provide software for instrument control and discuss how users can tune imaging parameters to accommodate diverse sample types. Thus, this protocol will serve not only as a guide for both introductory and advanced users adopting ASLM, but as a useful resource for any individual interested in deploying custom imaging technology.
Files
Files
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