Peripheral blood T colony-forming cells (T-CFC) from patients with T-cell malignancies can proliferate in methylcellulose in the absence of added growth factors or mitogenic stimulation. Mononuclear cells (MNC) from 7 patients with T-cell acute lymphoblastic leukemia were separated into cells forming rosettes with sheep erythrocytes (E+) or not (E-). E- cells were further depleted by complement-mediated cytotoxicity with OKT3 monoclonal antibody (E-OKT3- cells). The study of their spontaneous T-cell colony-forming ability suggested that proliferation of T-CFC in the absence of added growth factors requires cellular cooperation because: (1) No colony growth was observed at low cell concentrations (up to 2 X 10(4) cells/ml) whereas at higher cell densities the number of colonies increased exponentially; (2) The plating efficiency from unfractionated MNC was higher than that from E-OKT3- or E+ cells. Irradiated autologous E+ cells enhanced the plating efficiency from blast-enriched cell fractions (E-OKT3-) when co-cultured either directly in methylcellulose or separately in a two layer assay (agar-methylcellulose), suggesting that their activity could be due to diffusible factors; (3) Adherent-cell depletion of MNC decreased colony formation. Autologous irradiated adherent cells were able to restore the plating efficiency from MNCA- cells when co-cultured directly in methylcellulose but not in separate layers; however, media conditioned by patients' A+ cells could enhance the colony growth from patients' MNCA- cells, indicating that their activity could also be mediated by constitutively released soluble factors.