A. GHR-expressing Hek293 cells were pretreated with MβCD for 1 h, incubated with Cy3-GH for 30 min at 37°C and the fluorescence was visualised with a confocal microscope. Bar, 5 µm. B. Cells were pretreated as in Fig. 4A, followed by incubation for 15 min with GH at 37°C as indicated. Total cell lysates (TCL, upper panel) and GHR immunoprecipitations (lower panel) were analysed on western blot with the indicated antibodies. 130, mature GHR; 110, precursor GHR. C. Cells were silenced either for clathrin or for GFP (as a negative control), followed by incubation for 15 min with GH at 37°C as indicated. Total cell lysates (TCL, upper panel) and GHR immunoprecipitations (IP, lower panel) were analysed on western blot with the indicated antibodies. 130, mature GHR; 110, precursor GHR. Data are representative for two independent experiments (B).