Immunofluorescence staining
GGCX -/- cells were seeded on gelatin coated
cover-slips and transfected with vectors containing GGCX wt/mut .
After 24 hours of transfection cells were washed with PBS, fixed with
4% paraformaldehyde for 10 minutes and then blocked for 30 minutes with
10% FBS. Cells were incubated overnight with primary antibodies against
GGCX (Abcam, ab170921) and against PDI for ER staining (ThermoFisher,
MA3-019). Secondary Alexa Fluor 488 or 594-conjugated antibodies were
incubated for 1 hour at RT in dark. Mounting was performed with ProLong
Glass (ThermoFisher) including NucBlue. Images were taken by Apotome2
(Zeiss) and Zen software was used to calculated Pearson“s correlation
coefficient (PCC).