The deficiency of γ-glutamyl transpeptidase activity, which was evident in some but not all cystic fibrosis amniotic fluids, could be corrected by mixing with either normal fluids or nondeficient cystic fibrosis fluids. Incubation of any amniotic fluid for 20 min at 62° C resulted in total loss of γ-glutamyl transpeptidase activity, but the activity could be restored by mixing with untreated nondeficient fluids. In contrast, no restoration could be obtained by mixing with untreated deficient cystic fibrosis fluids. Dialysis of amniotic fluids did not diminish their corrective capacity. Only the transpeptidation reaction was corrected and no correction was observed for the hydrolysis or autotranspeptidation of γ-glutamyl p-nitroanilide in the absence of the glycylglycine or methionine acceptor. Plasma specimens did not have any corrective activity, although their γ-glutamyl transpeptidase activity could be restored after heat inactivation by mixing with untreated nondeficient amniotic fluids. No correction was found for aminopeptidase or disaccharidase activities. These findings suggest that the deficient cystic fibrosis amniotic fluids probably contain normal quantities of the γ-glutamyl transpeptidase enzyme but lack a heat-labile nondialyzable activator that is necessary for its transpeptidation catalytic performance. An assay for this transpeptidase activator may provide a valuable approach to identify at least a subgroup of cystic fibrosis patients.