Fig. 5

Long-term observation of the bioprocesses sensitive to phototoxicity via PRS-SIM under low excitation power. a TIRF-SIM imaging of clathrin coated pits (CCPs) over 5000 frames (Supplementary Video 1). Although the collected fluorescence was ~ 20-fold lower than those used for acquiring artifact-free GT-SIM image, PRS-SIM image still conveys high-fidelity ring-like structure and prevents most artifacts fulfilled in conventional SIM image. b Spatial distribution of CCP nucleation events across the plasma membrane of a SUM-159 cell over the whole imaging duration. c z-score of CCP nucleation calculated from 7 cells rapidly increases as extending the observation window. z-score gets larger than 4.95 when observation window is longer than 4 min, indicating that there is a less than 1% likelihood that the clustered pattern of CCPs’ nucleation could be the result of random occurrence. d Histogram of mean square displacement (MSD) of 3572 CCP tracks from 3 cells. e Dual-color time-lapse imaging of CCPs (green) and F-actin (red) in a live SUM159 cell during the adhesion process (Supplementary Video 2). The whole imaging duration is ~ 8 min and representative PRS-SIM denoised frames are displayed. f Zoom-in visualization of the interaction between CCPs and F-actin. The SR images (left) and the segmentation result of F-actin (right) are displayed. g Mander’s overlapped coefficient (MOC) of the CCPs referring to F-actin during the cell adhesion. Lower MOC values indicated most CCPs are located in the gap of F-actin filament. Two curves are calculated based on the segmentation results from conv. SIM (blue) and PRS-SIM (red) images, respectively. Scale bar, 0.5 μm (a), 5 μm (b, e), 1 μm (f)