Selective protein kinase C (PKC) activators and inhibitors and a physiological agonist, fMLP, were used to study superoxide production and PKC isoenzyme activation in human neutrophils. The data show that the classical PKC isoenzymes, α and β, were activated by TPA and at a time prior to NADPH oxidase complex assembly. fMLP induced activation of PKC-β over a similar time course. Inhibition of c-PKCs reduced, but did not block, TPA-induced superoxide production completely, suggesting additional PKC isoenzymes were involved beyond NADPH oxidase assembly. PKC inhibitors were unable to inhibit fMLP-induced superoxide generation, indicative of signal redundancy in the induction of superoxide generation in human neutrophils.