Peritoneal inflammation induced by sterile irritants led to accumulation of macrophages that were more responsive to lymphokines than macrophages from resident cell populations of untreated mice. Lymphokine responsiveness was quantitated by measurement of macrophage-mediated tumor cytotoxicity induced by supernatants from antigen-stimulated immune spleen cell cultures. Tumor cytotoxicity by lymphokine-activated inflammatory macrophages was about 10-fold greater than that by equal numbers of lymphokine-treated resident cells. Analysis of the time course of activation and of lymphokine dose-response demonstrated that the increased responsiveness of inflammatory cells was a consequence of quantitative changes. Lymphokine-responsive cells in both resident and inflammatory populations were identical; the numbers of responsive cells increased with inflammation. Changes in numbers of lymphokine-responsive cells during acute inflammation were coincident with similar changes in numbers of peroxidase-positive macrophages. This finding suggested that the increased lymphokine responsiveness of inflammatory cells was dependent upon influx of young peroxidase-positive mononuclear phagocytes rather than stimulation of resident macrophages. This concept was further strengthened by the finding that whole body x-irradiation diminished the numbers of both peroxidase-positive and of lymphokine-responsive macrophages in inflammatory and resident cell populations. These data suggest that the lymphokine-responsive precursor for the activated tumoricidal macrophage during both steady-state and acute inflammatory conditions was the newly formed blood-derived mononuclear phagocyte. The 10-fold increase in numbers of lymphokine responsive macrophages in peritoneal exudate over resident cell populations reflected a similar increase in macrophage turnover induced by inflammation.