Briefly, cells were washed thrice with PBS and fixed using 4% PFA in 1 × PBS for 12 min and then washed three times in PBS. For tissue analysis, fixed floating sections maintained in a cryoprotectant solution were washed in PBS before being immunostained. Fixed cells/tissues were blocked and permeabilized for 1 h at RT with 3% BSA/5% appropriate serum/1 × PBS in the presence of 0.1% Triton X-100 and 0.3 M glycine. Subsequently, cells/tissue sections were incubated with the indicated primary antibody either for 1 h at room temperature or overnight at 4 °C. Cells/tissue sections were then washed thrice with 1 × PBS and incubated for 1 h at RT with the respective secondary antibodies and 20 min with DAPI. Cells/tissue sections were washed thrice with 1 × PBS before analysis. Cells/tissue sections were analysed by confocal microscopy. Confocal image acquisition was performed using a Zeiss LSM 780 or LSM 710 laser scanning microscope (Carl Zeiss).
