6B2-B8 T cell hybridoma cells were used to immunize mice, and immune spleen cells were fused with NS/1 myeloma cells. One clone, designated RTH-7, reacted with 89.5% of rat thymocytes, 30.2% of rat spleen cells, and 42.3% of rat lymph node cells. The RTH-7 reacted with a subset of rat T cells but not with B cells. Double staining analysis demonstrated that RTH-7 stained a rat T cell subset distinct from R1-10B5-positive cells that were known to be equivalent to mouse Lyt-2. It was revealed that RTH-7 and W3/25 recognize different antigenic epitopes on the same molecule. The RTH-7 as well as W3/25 substantially inhibited the production of interleukin 2 by cells in mixed lymphocyte reaction and the lymphocyte proliferation induced by mixed lymphocyte reaction. The RTH-7 inhibited the lymphocyte proliferation induced by Con A whereas W3/25 failed to do so. The RTH-7 defined antigen has a molecular weight of 53,000 under reducing condition and 47,000 under nonreducing condition. The RTH-7 defined antigen showed a wide range of heterogeneity in pI (6.2-8.8). The associated molecule of approximate molecular weight of 27,000 was occasionally detected with the RTH-7 defined antigen in 6B2-B8 T cell hybridoma cells as well as peripheral T cells but not in thymocytes. Thus, RTH-7 detects a cell surface antigen of a functional T cell subset of rat origin.