A new mechanism for regulation of delayed-type hypersensitivity (DTH) was investigated. The subcutaneous injection without adjuvant of syngeneic epidermal Langerhans' cells (LC) pulsed with keyhole limpet hemocyanin (KLH) into BALB/c mice gave rise to DTH upon challenge of the ear with the same antigen. When such cells were transferred intravenously, DTH did not occur, although the titer of anti-KLH antibodies was high. Peritoneal exudate macrophages (M phi) pulsed with KLH gave rise to neither DTH nor antibody production. The intravenous transfer of KLH-pulsed LC into mice immunized subcutaneously with KLH in complete Freund's adjuvant at the same time (in the sensitization phase) had a suppressive effect on DTH in an H-2-restricted way. M phi did not have immunoregulatory effects. When radiolabeled LC and M phi were transferred intravenously, they migrated into the spleen, but when they were transferred subcutaneously, they stayed in the skin or migrated into the lymph nodes. In splenectomized mice immunized with KLH, the intravenous transfer of LC pulsed with KLH neither caused the production of anti-KLH antibodies nor suppressed DTH. When Ia was expressed on the surface of M phi, the cells could present antigens, as LC could. These findings suggest that the anatomic sites at which an antigen is presented (i.e., spleen or draining lymph nodes) rather than the kind of cell that first presents an antigen to the immune system is important in deciding whether the immune response that takes place is DTH or antibody production.