Depletion of macrophages from human peripheral blood mononuclear cells (PBMC) caused a marked decrease in galactose oxidase and sodium periodate, but not a calcium ionophore, stimulated Interferon-γ (IFN-γ) production. Reconstitution of such depleted cultures with galactose oxidase treated macrophages, but not lymphocytes, restored IFN-γ levels to those of control nonfractionated PBMC. Thus, galactose oxidase seemed to act on macrophages which in turn stimulated lymphocyte production of IFN-γ. Unlike human cells which have terminal galactose residues on glycoproteins, murine cell glycoproteins terminate their oligosaccharide component in the order N-acetyl-neuraminic acid followed by d-galactose, N-acetyl-glucosamine, and glycoprotein. Galactose oxidase or sodium periodate only activated murine macrophages to stimulate lymphocyte IFN-γ production after exposing d-galactose residues by the removal of the terminal N-acetyl-neuraminic acid residues with neuraminidase. Removal of such exposed terminal galactose residues with β-galactosidase inhibited the effect of galactose oxidase on murine macrophages. Taken together, these results strongly suggest that oxidation of terminal galactose residues on macrophages is the initial site of action of galactose oxidase and sodium periodate. Studies with Boyden chambers have shown that galactose oxidase-treated macrophages released a soluble factor which stimulates lymphocyte production of IFN-γ. Based on these findings, it appears that the oxidation of terminal galactose residues on the surface of macrophages leads to the induction and transmission of a soluble signal for lymphocyte production Of IFN-γ. © 1985.