Prolongation of rat pancreatic islet allograft survival by a prior 7-day period of tissue culture was demonstrated, confirming previous reports by others. We then sought to identity those cells in islets capable of stimulating allograft rejection (Ia antigen-bearing cells) and to determine whether such cells and/or their Ia antigens might be reduced by tissue culture. Freshly isolated and 7-day-cultured Wistar-Furth rat islets were incubated with a mouse anti-rat Ia nonpolymorphic monoclonal antibody, then with peroxidase-conjugated goat anti-mouse antibody, and processed for electron microscopy. Peroxidase (Ia)-positive lymphocytes, macrophages, and capillary endothelial cells were identified in fresh but not in cultured islets. A radioligand assay, using 125I-protein A, revealed a 45% decrease in binding of Ia antibody to cultured compared with fresh islet cells. We conclude that Ia antigen-bearing lymphocytes, macrophages, and capillary endothelial cells in rat islets are reduced by tissue culture and that this may account, at least in part, for the decreased immunogenicity of cultured islet allografts.