These studies examined the effects of X radiation and interleukin 3 (IL-3), which is an effective cytokine for the generation of megakaryocytopoiesis from X-irradiated hematopoietic stem/progenitor cells, on the terminal process of human megakaryocytopoiesis and thrombopoiesis. Mature megakaryocytes were induced by culturing CD34 cells from normal human peripheral blood in a serum-free liquid culture stimulated with thrombopoietin. The experiments contained the following groups: control cultures with nonirradiated cells incubated for 15 days; cultures treated with IL-3 on day 7 or day 11, cultures irradiated with 2 Gy on day 7 or day 11, and cultures treated with IL-3 immediately after X irradiation. The nonirradiated control cultures produced megakaryocytes from day 7, and both the megakaryocyte and platelet generation reached a peak on day 12–13. When X irradiation was performed on day 7, both the megakaryocyte and platelet numbers decreased remarkably, while no significant effect was observed on those numbers when cultures were X-irradiated on day 11. IL-3 showed neither protective nor promoting effects on the terminal stages of megakaryocytic maturation and platelet production. The results demonstrated that mature megakaryocytes are radiosensitive but that the radiosensitivity decreased with the terminal stages of megakaryocytic maturation, especially for the megakaryocytes entering into proplatelet formation.