The response of cells of the murine megakaryocytic lineage to human interleukin 6 (IL-6) was assessed in serum-depleted cultures using a variety of biological assays. IL-6 alone had no influence on megakaryocytic colony formation but augmented the numbers of these colonies induced by the multipotent colony-stimulating factor interleukin 3. However, in liquid marrow cultures, IL-6 alone promoted marked increments in megakaryocytic size and the activity of acetyl-cholinesterase, a marker enzyme of the lineage. Moreover, IL-6 induced a significant shift toward higher ploidy classes when megakaryocytic DNA was quantitated by flow cytometry. To determine whether the influence of IL-6 on megakaryocytic maturation was direct, the factor was added to cultures of single megakaryocytes isolated from megakaryocytic colonies. Fifty-four percent of these cells increased in size compared with 19% of those grown without IL-6. The data show that human IL-6 is a potent direct-acting growth factor for murine megakaryocytes with activity promoting maturation of that lineage.