Intrathymic (IT) antigen injection has been shown to induce antigen- specific systemic tolerance in the rodent. To delineate the mechanisms responsible for the induction of tolerance, we used the 2C line of T cell receptor transgenic mice. The majority of T cells in 2C mice express an antigen receptor specific for the major histocompatibility complex class I alloantigen L(d) and can be identified with the clonotypic monoclonal antibody 1B2. IT injection of lymphoid cells expressing L(d) was found to induce a significant prolongation in BALB/c skin allograft Survival. The allograft prolongation was associated with a marked reduction in the number of developing 1B2+ thymocytes (clonal deletion), which occurred primarily at the CD4+CD8+ stage of thymocyte development, as well as a reduction in the number of mature CD8+1B2+ 2C T cells in peripheral lymphoid tissue. In addition, CD8+1B2+ 2C T cells that survive deletion have decreased CD8 expression levels and a significantly reduced in vitro proliferative response to specific alloantigen (clonal allergy). Exogenous recombinant interleukin 2 restores the capacity of 2C T cells to respond in vitro to alloantigen. Experiments involving separation of cells by fluorescence-activated cell sorter indicate that there is a precise correlation between the reduction in CD8 expression and anergy induction. Collectively, these data indicate that IT antigen injection can induce antigen-specific systemic tolerance by both clonal deletion and clonal anergy.