Chlamydia trachomatis and Neisseria gonorrhoeae are the two most prevalent bacterial sexually transmitted infections reported in the United States. The purpose of this article is to review the various tests available for diagnosing C. trachomatis and N. gonorrhoeae. The nucleic acid amplification tests are preferred owing to their high sensitivity and specificity and use on specimens obtained noninvasively. Non-amplified non-culture tests, such as the DNA probe test, remain a choice for settings in which nucleic acid amplification tests are not available or affordable. Culture is recommended for use on conjunctival, oropharyngeal, and rectal specimens and in cases of suspected abuse. In addition, for gonorrhea, culture is recommended when screening in populations with a low prevalence of infection. Point-of-care tests are limited by low sensitivity and should be used for only populations unlikely to return for follow-up.