Cell fixation for imaging
Cells were fixed in a final concentration of 4% paraformaldehyde in 10
mM Tris-HCl buffer, pH 7.4 (stock: 16% paraformaldehyde solution in
distilled water, EM grade, 15710, Electron Microscopy Science) for 10-30
minutes at room temperature. They were then washed repeatedly in PBS to
reduce the background autofluorescence, before staining with DAPI
(4′,6-diamidino-2-phenylindole) at final concentration 50 ng
ml-1 for 10 minutes for nuclear staining, and/or
staining with antibodies for immunofluorescence. Images were captured by
using a Carl Zeiss Microscope stand Axio Observer 7 system or Marianas
Yokogawa-type spinning disc inverted confocal microscope.