Clonogenic cells from peripheral blood of 13/16 patients with T-cell malignancies generated colonies in methylcellulose in absence of added growth factors or mitogenic stimulation. Spontaneous colonies were also obtained from purified cell fractions (E-OKT3- and/or E+ cells) in 73% and 57% of the patients, respectively. No spontaneous colony growth was observed with mononuclear cells of patients with solid tumors, non-T-cell leukemias or normal subjects. Colonies consisted of acid-phosphatase-positive, myeloperoxidase-and PAS-negative lymphoblasts bearing T-cell surface markers. Although the phenotype of pooled colony cells from either unfractionated mononuclear cells or E-OKT3- -derived colonies varied from patient to patient, the colonies, like fresh leukemic cells, were mostly composed of relatively immature cells as assessed by the high proportion (greater than 40%) of OKT6+ and OKT10+ cells and the low proportion (less than 40%) of OKT3+ and/or E+ cells. Cytogenetic analysis of colony cells revealed either normal metaphases or chromosome anomalies similar to those observed in fresh leukemic cells. Moreover, cells from primary colonies exhibited a capacity for self-renewal in the absence of added growth factors.