HEK-293 cells transiently transfected with GPR30 (GPER1) were grown on poly-L-lysine-coated coverslips overnight. After 24 h, cells were fixed with 4% paraformaldehyde and 0.2% picric acid in phosphate buffer (pH 6.9) for 30 min at room temperature and washed several times with phosphate buffer. Specimens were permeabilized with ice-cold methanol and then incubated with Anti-GPR30 (GPER1) Recombinant Rabbit Monoclonal Antibody (Product # 703480, 1:500 dilution). Cells were then incubated with Alexa488-conjugated secondary antibody for 2 h at room temperature, mounted and examined using an immunofluorescence microscope. Note, strong staining of transfected cells.