Janus plasmonic-magnetic gold-iron oxide nanoparticles as contrast agents for multimodal imaging

1.2 Janus magnetoplasmonic nanostars (JMNSs) ..................................................................... 3 2 Nanoparticle sizes ....................................................................................................... 6 3 X-ray photoelectron spectroscopy (XPS) ................................................................... 7 4 Fitting of magnetic plots ............................................................................................. 8 4.1 Non-interacting Super-paramagnetic model: fit of M(H) measurements .......................... 9


Nanoparticle sizes
To calculate the size of the particles TEM was used.Several images per nanoparticle type were analyzed with the use of the software package Image J. The images were analyzed based on the gray scale contrast, making use of the threshold tools and particle analysis.
For the asymmetric nanodumbbells the gold part was first measured using its higher contrast (darker), and the iron oxide was them measured by removing the gold part and selecting the iron oxide based on grey scale levels.Manual measurements were performed where the grey scale did not offer a good contrast.The sizes of gold and iron oxide were calculated from the areas assuming a spherical shape (equivalent average diameter).For the JMNSs, only the total size was measured.Two different values were acquired: an equivalent average diameter, considering the area and assuming an equivalent spherical shape, and the Feret's maximum diameter that corresponds to the maximum tip to tip distance of the 2D projected nanostars.Error bars correspond to ± the standard deviation after the measurement of a minimum of 100 nanoparticles.
Average sizes are shown in the table and represented in the next graphs.
Ag with a full width at half maximum (FWHM) of 1.1 eV.The selected resolution for the spectra was 10 eV of pass energy and 0.15 eV/step.All measurements were made in an ultra high vacuum (UHV) chamber at a pressure around 5•10-8 mbar.The data was fitted using asymmetric and Gaussian-Lorentzian functions (after a Shirley background correction), where the FWHM of all the peaks were constrained while the peak positions and areas were set free.

Fitting of magnetic plots
The magnetic plots were analyzed by comparing the experimental results to several standard models. 3Below is the detailed description of the Langevin and non-interacting models used to fit the experimental results, and the plots after fitting the models a b

Non-interacting Super-paramagnetic model: fit of M(H) measurements
The standard Langevin approach to the superparamagnetism (ideal SPM model) provides quantitative information about the size of the particles. 4e magnetization  of a magnetic nanoparticle system as a function of external field  and temperature  is defined as: where  is the number of nanoparticles,  volume of a nanoparticle,  volume of the system,  o bulk saturation magnetization, and  the Langevin function. 5,6 the Langevin model the nanoparticle sizes are assumed to be Gauss distributed around the mean hydrodynamic diameter  h , and hence, the total magnetization of the system is not just , but an integral over the measured size distribution: The magnetization of saturation, [ s ] =• 2 (), is considered as a variable independent of the nanoparticles or domain magnetization, [] = ( 3 ) and both variables are related by the inorganic content of the sample and density.Often, domain magnetization initially  is fixed to the theoretical expected value, and with the fit a value for the mean size and polydispersity of the magnetic core is provided.Thus, the fit of these measurements at room temperature provides the calculation of the mean size and the standard deviation of the magnetic nanoparticles.

-Fit of ZFC/FC measurements
A non-interacting model was used to fit the obtained experimental measurements. 7The population of magnetic nanoparticles (given by a size distribution ) is divided in two groups at each temperature, depending on their nanoparticle size: the fraction in an ideal superparamagnetic state that corresponds to nanoparticles below a certain critical volume and those, above such limit, whose super spin remains blocked: In the first term, the low energy barrier approximation is used, where the energy barrier (defined as  eff , being  the nanoparticle volume) is much smaller than the thermal energy ( B  where  B is the Boltzmann Constant), and thus can be omitted.As a consequence, the response of the magnetization to changes of magnetic field or temperature ( or ) follows a Langevin function, where  is the particle magnetization (A/m in S.I.) and  s is the experimental saturation magnetization (including non-magnetic mass contribution, in general).The experimental magnetization and the nanoparticle magnetization are allowed to decrease with temperature following a spin wave-like behaviour 8 "Bloch type law" as: the Bloch constant () is obtained from the magnetization measurements as a function of temperature under the maximum field of 7 T, being between 2 and 4×10 -5 in all cases.
It is important to note that the anisotropy constant K is dependent of the temperature and it has to be taken into account to calculate the value. 9l the mentioned fittings are represented in Figure S6 (black curves).

CT attenuation pots
The experiments were performed starting with a highly concentrated solution, which was then successively diluted.The contrast efficiency was obtained by the slope after a regression line to a y = ax function.The graphs below represent the attenuation plots for every sample at three different energies.

Figure S3 :
Figure S3: TEM images of JMNSs with 16 nm iron oxide parts.

Figure S4 :
Figure S4: TEM images of JMNSs with 20 nm iron oxide parts.

Figure S6 :
Figure S6: XPS spectra of ND.20 sample.a) Fe 2p spectrum.The absence of satellite or shake-up peaks indicates that the sample is mainly Fe 3 O 4 .Both FeO and Fe 2 O 3 materials can be easily identified since they present satellite peaks at around 716 and 730, and 719 and 733 eV, respectively. 1,2b) Fe 3p fitted spectrum.The Fe 3+ /Fe 2+ theoretical ratio for pure Fe 3 O 4 is 2/1.The Fe 3+ /Fe 2+ ratio for the sample is around 2.2, which is in agreement with the shape of the Fe 2p spectrum.

Figure S9 :
Figure S9: Relaxation constants (1/T2 and 1/T1) as a function of iron concentration of the different JMNSs with 20 nm iron oxide part.The black lines show the linear fittings of the different plots.The slope (relaxivity) and the coefficient of determination are expressed for each fitting.

Figure S14 :
Figure S14: Control experiments showing a lack of Prussian blue staining (top) or scattered light (bottom), imaged using bright-field and dark-field microscopy respectively, after treatment of A549 cells with media alone, and thereafter stained and imaged using the same methods as for cells incubated with JMNSs.The lack of Prussian blue staining and scattered light confirms the specificity of both techniques for the visualization of iron oxide and gold respectively.

Table S1 :
Average diameter, measured by TEM, of the asymmetric nanodumbbells used as seeds in the JMNS synthesis.