Immunological aspects after orthotopic rat liver retransplantation (re-OLT) were examined in association with cell migration and mixed chimerism. At day 2 after the first orthotopic liver transplantation (day 0) in the combination of DA (MHC haplotype, RT1a) donor into PVG (RT1c) recipient, the grafted DA liver was removed and a new PVG liver was implanted into the same PVG recipient (re-OLT). In the PVG recipient at various times after the re-OLT, DA-derived antigen and cells were detected using a DA-specific anti-class I mAb R3/13 in conjunction with ELISA, immunoblotting, and immunohistochemistry. The level of soluble class I antigen, which had risen to 270 ng/ml after the first OLT, substantially decreased within 24 hr after re-OLT. Using immunoblotting, DA class I antigen was detected in the PVG recipient's lymphoid organs at day 3 after DA liver grafting and persisted for up to 21 days after the DA liver was replaced by a new PVG liver. Immunohistochemistry on sections of spleen from re-OLT rats showed that the level of migratory cells expressing DA class I correlated with the findings obtained by immunoblotting. While the DA-derived antigen and cells were detected in the re-OLT recipient, the DA-specific inhibition of mixed lymphocyte reaction was observed in re-OLT serum. Our results suggest that the implanted DA liver graft was the source of DA soluble class I antigen, but DA-derived antigen and cells detected in the re-OLT recipient organs could persist for a relatively long time under immunosuppression after the implanted DA liver was removed by re-OLT.