Immunofluorescence of wt or mutant αs. HEK293 cells stably expressing the β2-AR and HA-αs or HA-αs-R231H were incubated with (C, D, G, and H) or without (A, B, E, and F) 10 μM isoproterenol for 20 min and then fixed with 4% formaldehyde. αs was detected by incubation with mAb 12CA5, followed by fluorescein isothiocyanate-conjugated anti-mouse antibody (15). The β2-AR was detected by incubation with a polyclonal rabbit antiserum against its C terminus (14), followed by Texas red-conjugated anti-rabbit antibody (15). Images were visualized as described (14, 15).