Four monoclonal antibodies directed against human DR antigens were examined for their effects on lymphocyte activation in vitro. Two (DA6.121 and DA6.231) were serologically and biochemically indistinguishable and recognized a common determinant on the DR beta chain. One, DA6.164, reacted with an epitope probably on the beta chain of a subset of DR molecules while the fourth, DA6.147 was directed against an alpha chain determinant. All four produced measurable inhibition of lymphocyte activation. Responses to PHA or PWM were generally reduced by less than 30% and those to autochthonous or to allogeneic lymphoid cells by more than 30% some being completely abolished. DA6.121 and 231 were the most consistently effective antibodies and behaved in an identical manner. There was no additive effect when different monoclonal reagents were mixed. Reduction in the level of activation occurred when the responding population consisted almost entirely of T lymphocytes, suggesting that ADCC was not involved. The presence or absence of complement had a profound effect on the level of lymphocyte mitogenesis but responses were depressed by anti-DR monoclonals under both conditions. The results indicate that both the alpha and the beta chain of DR play some part in lymphocyte activation. It is suggested that the continuous regeneration of DR antigens on the surface of stimulator cells may be important in eliciting MLR.