Populations of lymphoid cells were obtained from the gastrointestinal tract of guinea pigs and compared with splenic cells for effector cell activity in assays of spontaneous cell-mediated cytotoxicity (SCMC), mitogen-induced cellular cytotoxicity (MICC), and antibody-dependent cellular cytotoxicity (ADCC). Lymphoid cells were isolated from mesenteric lymph nodes (MLN), Peyer's patches (PP), and the small intestinal mucosa. Mucosal lymphoid cells were separated into a population of cells containing intraepithelial lymphocytes (IEL), and a second population of cells containing intraepithelial lymphocytes (IEL), and a second population of cells containing lamina propria (LP) lymphocytes by a combination of physical and enzymatic methods. All populations of lymphoid cells from the gastrointestinal tract were capable of mediating MICC. Relative to spleen cells, IEL possessed more MICC effector cell activity whereas cells from MLN, PP, and LP possessed less activity. In contrast, SCMC and ADCC effector cell activities were present only in the IEL population and these cytotoxic activities were comparable to that of splenic cells. These results demonstrate that mucosal lymphoid cells possess cytotoxic effector cell activities that may contribute to local mucosal immunologic reactivity.