The relative value of culture, direct specimen antigen detection tests, i.e., enzyme-linked immunosorbent assay (ELISA) and immunofluorescence (IF) tests in the diagnosis of Chlamydia trachomatis infection was studied in 125 newborns and 121 adults with signs of conjunctivitis. Eye and nasopharyngeal samples were tested by culture using cycloheximide-treated or irradiated McCoy cells, ELISA (i.e., Chlamydiazyme, Abbott) and IF tests (i.e., Chlamyset, Orion and MicroTrak, Syva). Of the neonates, 70 (35 boys and 35 girls) and 54 (33 males and 21 females) of the adults were positive in one or both eyes in one or more tests: 191 (39%) in cultures, 173 (35%) in ELISA and 160 (33%), 176 (36%) in each of the IF tests. Using culture as standard reference, the sensitivities of ELISA and the IF tests were 88%, 81% and 87%, while the corresponding specificities were 99%, 98% and 97%, respectively. The predictive values for a negative test (PVN) were 93%, 89% and 92% and for a positive test (PVP) 98%, 96% and 94%. Of the 124 cases chlamydia-positive in the eyes, 67 (54%), 76 (61%), 64 (52%) and 70 (57%) were positive in nasopharyngeal samples in one or more of culture, ELISA and the two IF tests, respectively. The sensitivities of ELISA and the IF tests in nasopharyngeal samples were 87%, 78% and 81%, while the corresponding specificities were 90%, 93% and 91%, respectively. The predictive values for a negative (PVN) test were 95%, 92% and 93%, and for a positive test (PVP) 76%, 81%, and 77%. Nasopharyngeal swabs were more often positive in cases with 2 or more weeks' duration of symptoms than in those with shorter duration.