Supplementary Figure 8: STED microscopy of TCRβ or CD3ε and image autocorrelation analysis.
From: TCRs are randomly distributed on the plasma membrane of resting antigen-experienced T cells
Representative STED microscopy images of fixed primary murine CD4+ TEFF cells labeled with (a) KT3-scFv-AS635P (n = 2 biologically independent samples for both activating and non-activating conditions) or (b) H57-scFv-AS635P during interaction with non-activating or activating supported lipid bilayers as indicated (n = 5 and n = 1 biologically independent samples for non-activating and for activating conditions, respectively); Recorded images (left) were analyzed by image autocorrelation (right) and compared to simulated random distributions at molecular densities matched to the experimental data. Red curves represent simulated random data (mean ± SEM.; n = 5); blue curves represent analysis of multiple ROIs in the respective images (mean ± SEM.; n = 5). Red boxes in STED images indicate ROI positions chosen for ACF analysis. ROI positions were chosen similarly in other images (not shown). Scale bars: 3 μm; Cartoons illustrate the labeling strategy
