Effect of neutralizing anti-TPO antibody on rhIL-11– and IL-3–induced human MK colony formation. CD34⁺ cells were isolated from human BM and assayed in serum-free media for BFU-MK–derived (A) and CFU-MK–derived (B) colonies. The following cytokine concentrations were used: rhIL-11 (50 ng/mL), rhIL-3 (10 ng/mL), and rhTpo (10 ng/mL). Both the anti-Tpo antibody and isotype antibody were added into the cultures at 75 μg/mL. Suboptimal concentrations of growth factors and optimal concentrations of antibody were used to guarantee antibody excess. Clusters of 3 or more CD41⁺ cells were scored as a CFU-MK–derived colony, whereas multifoci and greater than 100 fluorescent cells were scored as a BFU-MK–derived colony. Each column with bar corresponds to the mean ± SEM of triplicate cultures from one experiment representative of three separate experiments.