Rectal and pharyngeal infections with gonorrhea and chlamydia are of concern because they are associated with higher risk of HIV acquisition. Extragenital screening in asymptomatic persons at high risk may have the potential to reduce the incidence of these sexually transmitted infections (STIs). Several testing platforms are available for the testing of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) using nucleic acid amplification tests (NAATs). Self-collected extragenital samples are currently not approved by the US Food and Drug Administration in any NAAT platform. This study compares the analytical performance of self-collected extragenital specimens to that of clinician-collected specimens. We performed a multicenter/multiplatform validation study as a National Veterans Health Administration Pathology and Laboratory Medicine quality improvement project, with 9 different participating sites. Self-collected specimens were obtained at the same time as clinician-collected specimens. Clinician-collected specimens were used as the gold standard to evaluate the sensitivity and specificity of self-collection. A total of 2324 individual tests were analyzed (501 rectal and 661 oropharyngeal). The sensitivity was 94.44% for CT and 100% for NG for rectal specimens, whereas it was 100% for CT and 97.22% for NG for oral specimens. Specificity for oral specimens was 99.85% for CT and 99.36% for NG, whereas for rectal specimens, it was 99% for CT and NG. Self-collected specimens for extragenital CT/NG testing are highly sensitive and specific, with negative predictive values of 100%. Self-collection has the potential to overcome a major barrier for STI screening by providing an accessible, convenient, and patient-centered alternative.