Analysis of β2AR trafficking in AAV-β2AR/EGFP infected HEK 293 cells. Cells were cultured in 10% FBS and treated with either vehicle (A), 10 μM isoproterenol for 20 minutes (B) or 10 μM isoproterenol for 24 hr (C) and analyzed via epifluorescence microscopy using polyclonal antibody to the cytoplasmic tail of β2AR. Mock infected HEK 293 cells demonstrated no β2AR staining (data not shown). Recombinant β2AR showed predominantly surface staining in the presence of vehicle (A). Following 20 minute isoproterenol treatment, recombinant β2AR were sequestered internally (B). Following 24 hour isoproterenol treatment, recombinant β2AR demonstrated trafficking to large, perinuclear vesicles with some β2AR demonstrated on the surface (C). This experiment was performed 3 times with identical results. Scale bar, 10 μM.