Staphylococcus aureus cells were injured thermally by exposure to 55°C for 15 min and allowed to recover for various lengths of time at 37°C in Trypticase Soy Broth. During recovery, thermostable nuclease (TNase) production was measured using a turbidimetric-spectrophotometric method. Production increased during recovery until approximately 2 h after injury when the amount of TNase began to decrease unexpectedly. Protease(s) was thought to be degrading the TNase, and positive results of gelatin agar diffusion tests and heat inactivation experiments supported this hypothesis. Protease inhibitor studies with ethylene diamine tetraacetate (EDTA) and phenyl methyl sulfonyl fluoride (PMSF) confirmed the involvement of protease(s) in the observed decrease in TNase activity. Implications of TNase inactivation in screening of foods for enterotoxigenic staphylococci are discussed.