Analysis of the global protein tyrosine phosphorylation in normal HPCs and AML cells. (A) The level of protein tyrosine phosphorylation in normal unstimulated and stimulated (GM-CSF, SCF, and IL-3 for 5 and 20 minutes or for 4 hours) CD34⁺ HPCs, fresh AML samples, and AML cell lines was studied under similar conditions by Western blot with the antiphosphotyrosine antibody 4G10. The membrane was stripped and reprobed for β-actin as a loading control (lower panel) and for Lyn (black stars). Results shown are representative of experiments performed with normal CD34⁺ HPCs from 3 healthy donors and 38 AML samples (patient nos. 1, 2, 4–22, 24–28, 30, 31, 33–37, 56–60). (B) A similar analysis was performed on fresh AML cells cultured alone or in the presence of PP2 (10 μM) for 1 hour. Results shown are representative of experiments performed with 20 AML samples (patients 1–20). Black stars indicate the position of Lyn.