Protein level, distribution, and activity of PTP-MEG2 in normal and PV ECFCs and the correlation with in vitro ECFC development. Normal and PV ECFCs were cultured in conditioned liquid medium and analyzed at the indicated times. (A-C) Whole-cell extracts (A) or cytosolic and membrane fractions (B-C) of day 6 (A-B) or day-8 to -21 (C) ECFCs from different PV patients and healthy volunteers were subjected to Western blot analyses with anti-PTP-MEG2 antibody. The far right lane in panel A represents extracts from normal day-6 ECFCs infected with retrovirus carrying PTP-MEG2 (Figure 4A). (D) Quantitative representation of the protein level of PTP-MEG2 (based on gel scanning) in cytosolic (open symbols) and membrane (closed symbols) fractions of day-8 to day-20 PV (▪) and normal (♦) ECFCs. Data represent the average of at least 4 experiments with different PV and normal samples. (E) PTP activity in the membrane extracts of day-8 to day-20 PV (▪) and normal (♦) ECFCs. (F) For erythroid colony-forming assays, 1000 day-8 to day-20 PV (▪) and normal (♦) ECFCs were seeded in semisolid methylcellulose medium under optimal culture conditions. Erythroid colonies were counted after 10 days of culture.