When eosinophil peroxidase (EPO) was incubated with Staphylococcus aureus (staph) approximately 40% remained firmly bound to the bacteria as indicated by guaiacol assay. The staph-EPO complex was killed readily by concentrations of H2O2 and a halide that did not alter the viability of control (non-EPO-coated) organisms or organisms pretreated with another basic protein, protamine. This toxic effect was abolished by omitting either H2O2 or halide and was inhibited by azide, cyanide, or aminotriazole. Peroxidase activity was evident on the surface of the organism after ingestion by rabbit blood monocytes. The staph-EPO complex was killed at a greater rate by both rabbit blood monocytes and alveolar macrophages than were control organisms, an effect that did not appear to be secondary to improved phagocytosis. The peroxidase inhibitor azide inhibited the killing of staph-EPO by these mononuclear phagocytes without affecting the killing of control staphylococci; in the presence azide, the killing curve with staph-EPO returned to that seen with control organisms. Further, aerosolized EPO-coated organisms were cleared more rapidly from rat lung than were uncoated organisms or organisms pretreated with protamine. These findings suggest mononuclear phagocytes can utilize EPO bound to the surface of ingested microorganisms to enhance their microbicidal activity.