Peyer's patches are thought to be deficient in macrophages capable of sustaining the induction of an immune response. The present study examined the position, morphology, and phagocytic activity of Peyer's-patch macrophages, and their ability to take up, in situ, a tracer antigen introduced through the intestinal lumen. Peyer's-patch macrophages were isolated in vitro, and nonspecific esterase activity of individual cells was quantitated. Macrophages concentrated within the epithelium overlying Peyer's patches and in the subepithelial zone showed phagocytic activity and the ability to take up exogenous ferritin. Adherent macrophages demonstrated heterogeneity of esterase activity, with approximatley 15% having none. However, esterase activity in macrophages from lymph nodes was equally heterogeneous. The profile of esterase activity of Peyer's-patch macrophages seemed intermediate between those of macrophages from submandibular and mesenteric nodes. Tingible body macrophages were concentrated in germinal centers and were poorly adherent. The results of this study indicate that macrophages similar to those in lymph nodes are concentrated in the region where they may intercept antigens entering through overlying specialized epithelium. The presence of plasma cells in the same region suggests that local reaction to antigens, perhaps modulating further antigen entry, is supported by these macrophages. Perhaps the proportion of functional macrophages in the entire Peyer's patch is too low to support immune induction in in vitro-tests.