A series of gray zone specimens with sample to cutoff ratio of 0.7 to 0.99 encountered in routine use of the Syva Microtrak (Syva, Inc.) Chlamydia trachomatis enzyme immunoassay (EIA) test for urogenital specimens were subjected to repeat testing in duplicate and high-speed centrifugation with direct fluorescent antibody testing of the centrifugate. Immunofluorescent C. trachomatis elementary bodies were observed in high-speed centrifugates in more than 40% of two series of gray zone specimens examined indicating that a C. trachomatis gray zone result would require confirmatory testing.