Functional analyses were performed on neutrophils isolated from 6 patients from two institutions who displayed features of chronic neutrophilic leukemia (CNL). These neutrophils demonstrated a consistent deficiency (44 ± 8% of control values) in superoxide anion (O-2) production in response to the phorbol ester, phorbol myristate acetate (PMA). O-2 production in response to chemotactic peptides was near normal (82.3 ± 10.7% of control values). Bacterial killing was normal in the two patients studied, and chemotaxis was diminished in response to zymosan-activated plasma and to high concentrations of chemotactic peptides in the patients studied. Cytosolic C kinase activity was decreased in one of the two patients studied. These results suggest that a deficient O-2 release in response to PMA is a hallmark of neutrophils in CNL and may provide a diagnostic indicator of this condition.