Systemic treatment with antibody to CD40 ligand (aCD40L) can prevent autoimmunity and transplant rejection in several animal models and is currently under evaluation in clinical trials. While it is known that aCD40L administration inhibits expansion and effector functions of aggressive T cells, it is still unclear whether additional regulatory mechanisms are operative. Here we demonstrate that a single episode of CD40L blockade during development of the autoaggressive immune response completely prevented autoimmune disease in the RIP-LCMV mouse model for virally induced type 1 diabetes. Interestingly, protection could be transferred by a highly potent, bitypic cell population sharing phenotypic and functional properties of both natural killer (NK) and dendritic cells (DC). Furthermore, protection of prediabetic recipients was autoantigen specific and did not result in generalized immunosuppression. The origin, function, and therapeutic potential of these bitypic NK/DC regulatory cells is discussed.