Despite increasing therapeutic use of interferon (IFN)-alpha, its effects on human neutrophil function are not well characterized. In vitro preincubation of neutrophils with recombinant IFN-alpha and -gamma, tumor necrosis factor (TNF), or granulocyte-macrophage colony-stimulating factor (GM-CSF) enhanced neutrophil respiratory burst responses to stimulation with influenza A virus (IAV) and FMLP. The enhancing effects of IFNs were more subtle and required more prolonged incubation than those of TNF and GM-CSF. TNF and GM-CSF enhanced neutrophil binding of IAV and neutrophil intracellular calcium and membrane depolarization responses to IAV or FMLP stimulation, while IFNs did not. Inhibitors of neutrophil tyrosine kinase activation and protein synthesis blocked IFN-alpha-induced enhancement of respiratory burst responses. In addition to its other well-characterized effects, IFN-alpha may protect against viral infection indirectly by promoting neutrophil respiratory burst responses.