The concentration of cytosolic ionized calcium, [Ca²⁺]_i, was measured in intact neutrophils by use of a fluorescent indicator trapped in the cytoplasm. A given rise of [Ca²⁺]_i elicited by the chemotactic peptide formylmethionylleucylphenylalanine (FMLP) was associated with a much greater degree of superoxide generation and myeloperoxidase secretion than was the same or larger [Ca²⁺]_i produced by a specific calcium ionophore, ionomycin, which bypasses cell surface receptors. Thus, FMLP appears to generate some important excitatory signal in addition to a rise in [Ca²⁺]_i, and exocytosis and superoxide generation in neutrophils may not be simply dependent on [Ca²⁺]_i as is widely supposed.