Antibodies recognizing MHC class I molecules expressed on the surface of T cell shave been shown ot inhibit T cell responses In vitro. These findings suggested that therapy with such an antibody may prevent rejection and promote graft an acceptance. We therefore tested the effect of an anti-HLA class 1 α3 domain antibody (TP25.99) in vivo transgenic C57BL/6 mice expressing HLA-B2705. Flow cytometric analysis confirmed the binding of Tp25,99 to normal human peripheral blood lymphocytes, and to mouse spleen cells, bone marrow cells and thymocytes isolated from hemizygous (+/-) transgenic littermates but not from homozygous (-/-) littermates. TP25.99 inhibited OKT-3-induced, but not PMA+ionomycin-induced, proliferation of human peripheral blood lyphocyte as well as anti-CD3 or Con-A-induced proliferation of HLA+ mouse T cells. Both intact monoclonal antibody TP25.99 and TP25.99 Fab inhibited T cell proliferation. Reduced proliferation was associated with suppressed production of interleukin-2 as measured by ELISA. The efficacy of TP25.99 Fab in vivo was evaluated in a heart allograft model. Antibody therapy of (H-2 b , B2705 + trasngenic recipients of allogeneic Balb/c (H-2 d ) heart grafts prolonged graft survival significantly (MST = 19.8 +/- 6.4, p = 0.003_compared to treated (H-2 b , B2705 - ) (MST = 9.17 +/- 2.2) or untreated (H-2 b , B2705 + ) (MST = 10.0 +/- 2.8) transgenic recipients. This demonstrates that immunomodulation through anti-HLA class I antibody therapy can lead to prolongation of graft survival.