Methotrexate, cyclosporin A and prednisolone have been shown to improve graft survival rates in solid organ transplantation. However, little is known concerning their capacity to promote lasting engraftment of allogeneic bone marrow. Therefore, we tested these agents in LEW rats receiving MHC-mismatched marrow after pretreatment with a myeloablative dose of busulfan plus different doses of total body irradiation (TBI). To avoid mortality due to graft-versus-host reaction (GHVR), F1(CAP x LEW) marrow was transferred. Hematological parameters were determined twice weekly to monitor engraftment and rejection. The pretreatment was lethal but not sufficiently immunosuppressive to ensure lasting engraftment in all animals. Thus, post-transplant immunosuppressive protocols could be evaluated for their capacity to improve engraftment rates. Standard clinical doses of methotrexate (0.25 mg/kg i.p. day 1, 3, 6, 11, 18, 25), cyclosporin A (10 mg/kg orally day 0-28) and prednisolone (1 mg/kg i.p. day 0-28) were administered and proved to be of nearly equivalent toxicity in our system. All three agents failed to allow engraftment after busulfan alone. After additional conditioning with 1.5 Gy of TBI, methotrexate and cyclosporin A reduced the rejection rate from 100% to 59% and 70%, respectively. When 3 Gy of TBI were added to busulfan, cyclosporin A and prednisolone were able to reduce the rejection rate from 67% to 33% and 39%, respectively, whereas 0.12 and 0.25 mg/kg methotrexate completely prevented graft rejections. After cessation of cyclosporin A therapy, late secondary rejections were frequently observed. These results demonstrate that postgrafting immunosuppression with protocols conventionally used for the prophylaxis of GVHR is able to facilitate lasting engraftment of MHC-mismatched bone marrow.(ABSTRACT TRUNCATED AT 250 WORDS)