Twelve human T-cell lines were derived from cultures stimulated with phytohaemagglutinin (PHA) or with the recall antigen purified protein derivative (PPD), using the one-step agar colony method, followed by further expansion of individual T-cell colonies for at least 50 days. As judged by monoclonal antibodies, two of the PHA-derived T-cell lines carried the helper-inducer (H-I) phenotype and two the suppressor-cytotoxic (S-C) phenotype. Four PPD-derived cell lines displayed the H-I phenotype, and three of these proliferated specifically when challenged with the recall antigen, whereas the fourth did not. Four PPD-induced T-cell lines carried the S-C phenotype, and only one of these was antigen-specific. All cell lines with H-I phenotype displayed helper cell activity as determined by Ig secretion of B lymphocytes stimulated with a T-cell-dependent polyclonal B-cell activator. Cell lines with the S-C phenotype had no such helper activity and suppressed Ig responses in the presence of freshly isolated T cells. Thus, a good correlation between phenotypic markers and functions was found. T helper cells and T suppressor cells can be differentiated by means of both polyclonal and antigenic stimuli.