We found that the cytosol of rat peritoneal polymorphonuclear neutrophils contains factor(s) that can stabilize an unstable enzyme, nitric oxide synthetase, in the cytosol. This enzyme has been purified to a single protein from the cytosol. Its half-life was 3 hours at 4 degrees C and was prolonged to greater than 24 hours by the stabilizing factor in the cytosol. The molecular weight of the stabilizing factor was greater than 100,000. Its activity was lost by the treatment with heating or alkali for 1 min or with acid for 5 min. It did not adhere to the carboxymethyl or diethylaminoethyl column at neutral pH. This stabilizing factor(s) may play a role in the regulation of the nitric oxide synthetase.