The present study was undertaken to elucidate the role of T cell subsets in regulating the in vitro growth of human granulopoietic progenitor cells (CFU-C). Prior to CFU-C assay, mononuclear cells obtained from bone marrow and cord blood were depleted of T cells or functionally distinct T cell subsets by the method of complement-mediated cytolysis with the use of monoclonal antibodies, OKT3, OKT4 and OKT8. The depletion of any T cell subset of OKT3+, OKT4+ or OKT8+ cells from bone marrow and cord blood cells showed no significant alterations in the generation of CFU-C. The supplementation to the in vitro culture system of OKT4+ or OKT8+ cells, which had been negatively selected by complement-mediated cytolysis using the mutually exclusive monoclonal OKT8 or OKT4 antibody, respectively, did not alter the growth of CFU-C-derived colonies without mitogenic stimulation. In contrast, the production of colony-stimulating activity (CSA) in peripheral blood mononuclear cells was significantly reduced by removing not only OKT3+ cells but also OKT4+ or OKT8+ cell subsets. There were no significant differences in the degree of reduction between the different procedures. These results suggested that T cell subsets played an important role in the regulation of steady-state granulopoiesis through the stimulation of CSA production. The presence of a specific subset of T cells in CSA production was not demonstrated.