We compared the performance of a commercial enzyme immunoassay (EIA) (Chlamydiazyme; Abbott Diagnostics, North Chicago, Ill.) with that of cell culture for the detection of Chlamydia trachomatis cervical infection in 1,417 women attending public health clinics. Confirmatory chlamydial testing by a direct fluorescent-antibody test (MicroTrak; Syva Co., Palo Alto, Calif.) was performed on specimens from women who had positive EIAs. Overall, only 57% of women who had a positive chlamydial test by cell culture were also positive by EIA. We noted a strong association between the number of chlamydial inclusions in cell culture and a positive EIA outcome. The proportion of culture-positive women who also had a positive EIA declined with age and a history of previous sexually transmitted disease and increased among oral contraceptive users. The results of direct fluorescent-antibody confirmatory testing suggested that cell culture was also insensitive for the detection of C. trachomatis infection. Our observations demonstrate that the performance of the chlamydial EIA may vary greatly with individual patient characteristics and that the utility of EIA as a screening test may be limited, especially in older women.