The next-generation amplification test for Chlamydia trachomatis and Neisseria gonorrhoeae (Roche cobas 4800), a fully automated system, was compared head to head, using female samples, to Gen-Probe Aptima Combo 2 and BD ProbeTec using Viper. Endocervical swabs, female urine, and endocervical samples in liquid-based cytology medium were run on at least two of three platforms. A total of 4,316 samples were evaluated, and 281 chlamydial and 69 gonococcal infections were identified. Estimates of sensitivity and specificity were obtained relative to the patient infection standard (PIS) and using latent class analysis (LCA). Chlamydia sensitivity estimates ranged from 86.9 to 95.6% using PIS and 97.6 to 98% using LCA. Specificity was ≥ 99.6% for all sample types. Sensitivity ranged from 95.6 to 100% using PIS and 96.9 to 100% using LCA for the detection of gonococcal infections. Specificity for gonococcal infections was ≥ 99.8%. cobas 4800 performance was equivalent to the comparator assays (all P values, >0.05), and the fully automated system provides high laboratory efficiency.