Here we report a new anti-rat Thy-1 monoclonal antibody (MAb) 12C5 that recognized a novel epitope on a GPI-anchored Thy-1 antigen. In rat thymocytes, MAb 12C5 reacted with 25 kDa Thy-1 antigen purified with MAb OX7. In rats, MAb 12C5 was expressed on 94% of thymocytes, 46% of spleen cells, 31% of mesenteric lymphnode cells, 76% of bone marrow cells, and 24% of splenic T cells. Furthermore, MAb 12C5 reacted with the hematopoietic cells in Mongolian gerbils, hamsters, and AKR mice (Thy-1.1) as in rats. In AKR mice, almost all bone marrow cells reacted with MAb 12C5. Thus, MAb 12C5 recognized a novel epitope of Thy-1 that was widespread in hematopoietic cells of rats and mice, as well as other rodents.