Reduced transgene expression in platelets of Yall;Mpl^−/− mice, but normal Tpo levels and platelet half-life. (A) Mpl mRNA (top) and protein (bottom) expression in purified platelets and megakaryocytes. qPCR was performed with primers amplifying both transgenic and endogenous Mpl; Itga2b (CD41) primers were used for normalization. n.d. indicates not determined. Note that Mpl mRNA in Yall;Mpl^−/− platelets is approximately 100-fold lower than in platelets of the other 2 genotypes. Similarly, Mpl protein is strongly decreased in platelets from Yall;Mpl^−/− mice. Membranes were reprobed with anti-CD41 antibodies to control for equal loading. (B) Tpo levels in mice with the indicated genotypes. Tpo concentration in both serum and plasma was determined by ELISA. Consistent with previous reports, Tpo is elevated in Mpl^−/− mice. In contrast, Yall;Mpl^−/− mice displayed slightly reduced Tpo compared with controls. Note that the plasma values are generally lower than the serum values, with the exception of Mpl^−/− mice. (C) Half-life of platelets is normal in Yall;Mpl^−/− mice. After injection of chemically activated biotin, blood samples were taken at indicated time points and the fraction of biotin-labeled platelets was determined by flow cytometry. Error bars in panels A through C represent SD.