Experiments were performed to test the hypothesis that stimulating antigens in the mixed lymphocyte reaction (MLR) are expressed in a restricted fashion on lymphocytes. Thus one-way MLR between spleen cells from congenic mouse strains differing at H-2 were treated with 5-bromo-2-deoxyuridine (BrdUrd) and light to inactivate dividing cells. The ability of the residual cells to act as responder or stimulator cells in a subsequent MLR was assessed. As expected, a specific loss of responder cell activity occurred under these conditions. Furthermore, such cells, following irradiation, would not stimulate fresh spleen cells from the strain against which they initially responded, but would stimulate spleen cells, following irradiation, would not stimulate fresh spleen cells from the strain against which they initially responded, but would stimulate spleen cells of unrelated mouse strains. This was shown to be due to the generation of cytotoxic T cells detectable in 51Cr release assays in the primary MLR, despite BrdUrd and light treatment and irradiation. Furthermore, loss of stimulator activity mapped to the H-2K region of the H-2 complex rather than the strong MLR-stimulating I-A or I-B subregions. These experiments provide no evidence of restricted expression of MLR-stimulating antigens on lymphocytes.