We have made a detailed analysis of the fate of Langerhans cell—free cultured keratinocyte allografts in two rat strain combinations, DA-to-PVG and DA-to-LEW, and compared the results with the rejection of conventional skin allografts in these strain combinations. The cultured keratinocyte layers were grafted both to the body surface using a technique to prevent wound contraction, and to the renal subcapsular site. Histological examination of grafts was made on days 2, 7, 10, 14, and 28 after transplantation. Donor-specific anti—class I MHC monoclonal antibodies were used to verify the donor origin of the keratinocytes. We report that the keratinocyte allografts are acutely rejected—but, in contrast to the conventional allografts, do not evoke alloantibody responses. Rejection of the keratinocytes at the renal subcapsular site was as rapid as that of conventional skin grafts. However, rejection of keratinocyte grafts on the body surface was delayed by a few days when compared with conventional skin grafts. Immunosuppression with cyclosporine prevented the rejection of DA keratinocyte layers placed at the renal subcapsular site of PVG rats, but rejection followed soon after cessation of cyclosporine therapy. These data sugest that rejection is a major constraint for the clinical application of cultured keratinocytes, and that autografts must be used if permanent cover is required. Moreover, the findings have interesting theoretical implications relating to the much greater vulnerability to rejection of skin grafts compared with organ grafts. Out current and previous data also suggest that class II-positive dendritic cells are the major stimulus to alloantibody production afte tissue transplantation.