Lyt-123/23 and Lyt-1 T cell subsets, positively selected by separation on the fluorescence-activated cell sorter, were tested in vitro for their role in the generation of H-2-restricted and alloreactive cytotoxic T cells. It is shown that in the proliferation assay, both T cell subsets responded equally well to H-2 and non-H-2 antigens (H-Y), respectively. In contrast, none of the selected Lyt subsets, but only the mixed population containing Lyt-1 and Lyt-123/23 lymphocytes gave rise to both H-2-restricted (anti H-Y, anti-trinitrophenyl) and alloreactive (anti-H-2) cytotoxic lymphocytes. The data imply an essential role of Lyt-1 cells as inducers or helpers in the generation of all cytotoxic lymphocytes from their precursors in the Lyt-123/23 pool.