Characteristics of Epo-induced GAB1 tyrosine phosphorylation. Serum- and growth factor-deprived UT-7 cells were stimulated for various times with 10 U/mL Epo (A) or for 10 minutes with various Epo concentrations (B). Cleared lysates were then prepared and immunoprecipitated with anti-GAB1 antibodies. Immunoprecipitates were analyzed by antiphosphotyrosine (anti-PY) antibodies. Assuming an equilibrium constant of dissociation of 200 pmol/L for the Epo receptor in UT-7 cells,29 receptor occupancy is 2% for 10 mU/mL Epo, 18% for 100 mU/mL, 70% for 1 U/mL, 95% for 10 U/mL, and 99.5% for 100 U/mL. Because apparent binding equilibrium was not achieved after 10 minutes of incubation for Epo concentrations less than 1 U/mL (data not shown), receptor occupancy for Epo concentrations of 10 mU/mL and 100 mU/mL was probably slightly lower than the indicated values.