1. The effects of antigen (Ag) injection on the distribution of lymphocyte populations of Cornell K-strain male chickens were studied. 2. Two experiments were conducted. In the first, chickens were injected with Brucella abortus (BA), a purported T-independent antigen. In the second, chickens were injected with sheep red blood cells (SRBC), a T-dependent antigen. Peripheral blood lymphocytes (PBL) and spleen lymphocytes isolated at 0, 3, 6, 9, 12 and 24 h following Ag injection were stained with monoclonal antibodies (mAb) detecting B-lymphocytes, CD4+ and CD8+ cells. 3. B-lymphocytes in the blood or spleen showed no significant changes following either BA or SRBC injection. In contrast, CD4+ cells were decreased in the blood and increased in the spleen following BA and SRBC injections. CD8+ cells were decreased in both blood and spleen following BA injection but were unchanged in either blood or the spleen following SRBC injection. 4. These results indicate that there is a change in both spleen and circulating lymphocyte populations, especially T-helper cells, following Ag injection. T-helper cells are apparently the primary population involved in the initiation of humoral immunity.