IL-10, which is secreted by multiple cell types, has regulatory effects on macrophages, including decreasing their ability to kill some microorganisms. The experiments presented here test the hypothesis that endogenous IL-10 inhibits the ability of macrophages to kill the facultative intracellular bacterium, Listeria monocytogenes. We show that the nonbactericidal macrophage hybrid, H36.12j (12j), can kill Listeria after incubation for 3 days with anti-IL-10 mAb. IL-10 was not detected in 12j macrophage supernatants by ELISA. However, flow cytometric analysis revealed high levels of IL-10 on the 12j cell surface. This indicates that macrophages that fail to secrete IL-10 may express IL-10 on the cell surface, and this IL-10 appears to suppress listericidal activity. Surface IL-10 is not unique to the 12j macrophage hybrid and may correlate with the absence of bactericidal activity in other macrophages. For instance, nonlistericidal resident and thioglycolate-elicited peritoneal exudate cells have 24 to 72% IL-10-positive macrophages. In contrast listericidal proteose peptone-elicited peritoneal exudate cells contained < 5% IL-10-positive macrophages. Whether this IL-10 is an integral membrane protein or is bound to IL-10 receptors is not yet known. However, the IL-10 does not elute with acid as other passively bound molecules do, nor does exogenous IL-10 bind to macrophages. In either case, since anti-IL-10 induces nonbactericidal macrophages to become bactericidal, the surface IL-10 is biologically active, and it appears to regulate macrophage bactericidal activity.