Addition of beta-lactamase to cultures of antibiotic-sensitive Escherichia coli protected the bacteria against lysis induced by either a hydrolyzable (cephalothin) or relatively nonhydrolyzable (ceftriaxone) cephalosporin. The later addition of a nonhydrolyzable, non-lysis-inducing beta-lactam antibiotic (oxacillin), which had a higher affinity for the beta-lactamase than ceftriaxone, allowed the reversal of the protection and the onset of lysis. These results suggest that trapping of the antibiotic by the enzyme, without significant hydrolysis, is a reversible process that may play a role in the resistance of some gram-negative bacteria to third-generation cephalosporins.