Trichomonas vaginalis PCR using reagents from a commercially available assay for Chlamydia trachomatis and Neisseria gonorrhoeae was evaluated for detection of infection in women and men attending a sexually transmitted disease clinic. Evaluations included three primer sets, endocervical swabs, vaginal swabs and urine, and various storage conditions. The TVK3/TVK7 primer set was optimal in our hands with sensitivities ranging from 69.5 to 96.8%. In all comparisons, T. vaginalis PCR performed better than routine diagnostics using microscopy for women and culture for men (P > 0.05). The assay performed well for all sample types tested, and vaginal swabs were stable for up to 7 days at ambient temperature. Using samples prepared for, and reagents from, the C. trachomatis-N. gonorrhoeae PCR assay allowed incorporation of T. vaginalis PCR diagnosis into routine clinical testing.