Fig. 2

Characterization of Atto 647N in live cells. a A co-localization study employing MitoTracker Green as the standard mitochondrial marker. Live U2OS cells were simultaneously stained with MitoTracker Green (green, 400 nM) and Atto 647N (magenta, 15 µM) for 30 min and imaged by SIM. b Enlargements of the boxed regions in a. c Time-lapse SIM images of live U2OS cells stained with MitoTracker Green (green, 400 nM) or Atto 647N (magenta, 15 μM). d The percentage of retained fluorescence intensity of Atto 647N and MitoTracker Green after 420 s of time-lapse SIM imaging (from n = 10 areas, mean ± s.d.; ^*** P < 0.0001, two-tailed t-test; statistics were performed using SPSS 19.0 software package (IBM Co.)). e Confocal images of U2OS cells stained with Rhodamine 123 (2 µM), MitoTracker Green (400 nM), or Atto 647N (15 µM) before and after cell fixation. Each SIM frame was acquired over 270 ms (i.e., a raw data exposure time of 30 ms). For the time-lapse images, consecutive SIM frames spaced at ~ 1.15-s intervals were obtained; representative images of consecutive SIM frames are displayed. Scale bars, a 5 μm, b, c 1 μm, and e 10 μm