Because few clinicians have access to laboratories offering cell culture confirmation of suspected Chlamydia trachomatis genital infections, we evaluated a diagnostic method in which fluorescein-conjugated monoclonal antibodies were used to directly identify C. trachomatis elementary bodies in slides made from genital secretions. Compared with culture results, the direct smear had a sensitivity of 92% and a specificity of 96% in 576 men, most of whom had symptoms and signs of urethritis. Among 595 women attending the same clinics, sensitivity of the direct smear for cervical infection was 89% and specificity was 99%. In 225 pregnant women screened in a prenatal or abortion clinic, the sensitivity and specificity of the test were 86% and 99% respectively. Direct detection of elementary bodies in genital smears offers an alternative diagnostic approach for C. trachomatis infections.