HeLa Kyoto cells (kindly provided by Belousov V.V., Shemyakin‐Ovchinnikov Institute of Bioorganic Chemistry, Moscow) were imaged 24–48 h after the transient lipofectamine transfection before and after 2.5 μm Ionomycin addition using a laser spinning disk Andor XDi Technology Revolution multi‐point confocal system (Andor Technology, Belfast, UK) as previously described [5 (link)].
For fluorescence recovery after photobleaching (FRAP) experiments, HeLa Kyoto cells were transfected as described above and imaged using a Leica SP5 STED confocal microscope (Leica‐Microsystems, Bensheim, Germany) and 70% of 488 nm laser power for bleaching (power at 100%—65 mW) during 1000 ms, with the capture settings: 100 ms per frame, 20 and 600 frames before and after bleaching, respectively, resolution—16 × 16 pixels, pixel size—0.38 μm as described earlier [5 (link)].
For fluorescence recovery after photobleaching (FRAP) experiments, HeLa Kyoto cells were transfected as described above and imaged using a Leica SP5 STED confocal microscope (Leica‐Microsystems, Bensheim, Germany) and 70% of 488 nm laser power for bleaching (power at 100%—65 mW) during 1000 ms, with the capture settings: 100 ms per frame, 20 and 600 frames before and after bleaching, respectively, resolution—16 × 16 pixels, pixel size—0.38 μm as described earlier [5 (link)].
Free full text: Click here
