Despite the fact that the Peyer's patch (PP) is the primary site for antigen uptake in the intestine, the cellular basis of antigen handling after transport into the PP is poorly understood. We performed immunohistology of routine PPs using the dendritic cell (DC)-reactive monoclonal antibodies N418, NLDC-145, M342, and 2A1, as well as antibodies to other T cell, B cell, and macrophage markers. N418 § 2A1 +, NLDC-145, M342cells form a dense layer of cells in the subepithelial dome (SED), just beneath the follicle epithelium, and are scattered throughout the follicle, sparing the germinal center. In contrast, N418 +, 2A1 +, NLDC-145 +, and M342 § DCs are present in the interfollicular T cell regions (IF[L). CD3 + and CD4 § but no CD8 + T cells were present in the SED and the follicle, including the germinal center, while CD3 +, CD4 +, and CDST ceils were present in the IF[L. B cells and macrophages were poorly represented in the SED as no B220 + cells, only few Mac-11~ cells, and no F4/80 + cells were present at this site. In contrast, Mac-1 h~ cells were found in the IF[L and lamina propria of intestinal villi, while F4/80 § cells were found only in the latter. In further phenotypic studies, we analyzed surface molecules of PP and spleen DCs by flow cytometry and found that these ceils had similar fluorescence profiles when stained with N418, NLDC-145, and 33D1 DC-reactive antibodies, and antibodies to the costimulatory molecules B7-1 (1G10) and B7-2 (GL1). In contrast, PP DCs expressed 5-10-fold higher levels of major histocompatibility complex class I! antigens ([E k) than spleen DCs. Finally, in functional studies, we demonstrated that both PP and spleen DCs process soluble protein antigens during overnight culture and induce similar levels of proliferation in CD3 § T cells, and CD4+/Mel 14 hi T cells from T cell receptor transgenic mice. The in vivo relevance of such presentation was shown by the fact that PP DCs isolated from Balb/c mice after being fed ovalbumin stimulated proliferation in ovalbumin T cell receptor T cells. Taken together, our data suggest that DCs in the SED of the PP are uniquely positioned for the processing of antigens passed into the PP from the overlying M cell, and that PP DCs are effective at processing and presenting oral antigens to naive T cells.