Fig. 3

SPLIT-FLCS of EGFP in the cytoplasm of live cells. a Representative filtered ACFs of EGFP in the cytoplasm of a HeLa cell at a STED power of 50 mW for different values of the parameter r ₁. Solid lines are a fit of the data to Eq. (15). Residuals are shown on the right axis. The image of a cell is shown in the inset (scale bar 10 μm). b Transit time t _D (solid squares) and apparent diffusion coefficient D (open squares) for different spatial scales, obtained as the average of single-point measurements in the cytoplasm of n = 12 different HeLa cells (mean ± s.d.). The solid line is a fit to Eq. (14) (D = 25 μm² s⁻¹). c Corrected value of the number of molecules N ^# vs. w ², for all the measured cells. The dashed lines are linear fits to the data. The slope varies from cell to cell according to differences in EGFP expression level. d Transit time t _D and e apparent diffusion coefficient D at different spatial scales for three different cells. f Transit time t _D and g apparent diffusion coefficient D as measured on the very same spot by three short (t _acq = 20 s) consecutive acquisitions