Sera from patients with aplastic anemia and amegakaryocytic thrombocytopenia contain an activity that stimulates megakaryocyte colony formation in vitro. We have assayed this megakaryocyte colony-stimulating activity (Meg-CSA) in sera of four patients receiving intensive antileukemic chemotherapy to determine whether the appearance of Meg-CSA is a physiologic response to the suppression of megakaryocytopoiesis. Three of the four patients were receiving consolidation or late intensification therapy for acute myoblastic leukemia (AML) in remission. The fourth was receiving induction therapy for de novo AML. During all or part of four chemotherapeutic cycles, serial Meg-CSA levels were assessed and correlated with the corresponding peripheral platelet counts. All courses of cytotoxic chemotherapy resulted in increases in serum Meg-CSA comparable to activity levels present in sera from patients with aplastic anemia. Two of the three patients studied during the early postchemotherapy interval manifested initial serum Meg-CSA elevations seven days before their thrombocytopenic nadirs when platelet counts were still between 100,000/mm3 and 140,000/mm3. Bone marrow recovery from chemotherapy was characterized by a decrease in serum Meg-CSA to pretherapy levels that occurred concurrently with the rise in platelet count to normal. These observations support the hypothesis that Meg-CSA is a physiologic humoral regulator of megakaryocytopoiesis elaborated in response to the depletion of either bone marrow megakaryocytes or megakaryocyte progenitor cells.