Active Thrombopoietin (TPO)

Thrombopoietin (TPO) is a humoral growth factor that is necessary for megakaryocyte proliferation and maturation, as well as for thrombopoiesis.TPO can promote platelet production, aggregation, ECM adhesion, and activation. It is principally produced in the liver and is bound and internalized by the receptor Tpo R/c-mpl. This protein is the ligand for MLP/C_MPL, the product of myeloproliferative leukemia virus oncogene. Mutations in this gene are the cause of thrombocythemia 1. Besides, GCSF has been identified as an interactor of TPO, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant mouse TPO and recombinant mouse GCSF. Briefly, biotin-linked TPO were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to GCSF-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30 min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µl stop solution to the wells and read at 450 nm immediately. The binding activity of TPO and GCSF was shown in Figure 1, the EC50 for this effect is 0.68 ug/mL.