Antibodies are produced when antigen-presenting cells (APC) pulsed with an antigen are injected intravenously (i.v.) into BALB/c mice, but subcutaneous (s.c.) injection of such APC causes delayed-type hypersensitivity (DTH). To identify the anatomic sites where T and B cells are activated, we labelled splenic dendritic cells (DC) with a fluorochrome, PKH 26, injected them i.v. or s.c., and used the label to locate them. When the DC were injected i.v., germinal centres in the spleen were hyperplastic on day 1. Most DC moved to T-dependent areas of the white and red pulp on day 1 and remained there at least until day 5, but no DC migrated into the lymph nodes. When the DC were injected s.c., they were in the sinus on day 1 and had entered T-dependent area of draining lymph nodes only by day 3; hyperplasia of germinal centres in the spleen and migration of DC into the spleen were not found. We used the polymerase chain reaction (PCR) to study which mice had spleen cells and lymph node cells that produced the cytokines interleukin (IL)-2, IL-4, IL-10, and interferon-gamma (IFN-gamma). In the sensitization phase, day 1 after DC injection i.v., almost all IL-10 transcript was found in spleen cells, but after DC injection s.c., IL-2 message was most abundant in lymph node cells. The expression of mRNA for IL-4 and IFN-gamma in mice that received DC i.v. was not different from that in mice that received DC s.c. in this phase. Immunohistochemical staining showed that cells stained for IL-10 were in the T-dependent area of the spleen from mice that received DC i.v. 1 day after the injection. Three days after the injection of DC i.v., cells stained for IL-10 were in the germinal centres as well. The number of such cells in the spleen of mice that received DC i.v. was significantly more than that in mice that received DC s.c. IL-10 may be important in development of TH2 response.