Growth in plasma clot culture of megakaryocyte colonies from adult bone marrow cells using 4 different erythropoietin (Ep) sources as the stimulating factor was reported. A major proportion of the megakaryocyte colonies (75%) were pure, while others were mixed, involving erythroblasts and megakaryocytes. Ultrastructural studies showed that megakaryocyte maturation was essentially normal and platelet shedding occurred. Megakaryocyte colony formation required many plated cells (greater than 3 .times. 105/ml). In the absence of erythropoietin, rare spontaneous megakaryocyte colonies could be observed, while no erythroid colonies were present. Erythropoietin induced a 5-fold increase in total colony number. Erythropoietin is apparently involved in human megakaryocyte differentiation but it does not act alone, since another factor related to seeded cell number appears essential for formation of human megakaryocyte colonies.