Given the many options available, selecting an HIV test for a particular clinical or research setting can be daunting. Making an informed decision requires an assessment of the likelihood of acute infection in the test population and an understanding of key aspects of the tests themselves. The ability of individual tests to reliably detect HIV infection depends on the target(s) being detected, when they can be expected to be present after infection, and the concentration of stable target in test specimens, all of which are explained by the virologic and serologic events after infection. The purpose of this article is to review the timeline of HIV infection, nomenclature, and characteristics of different tests; compare point-of-care and laboratory-based tests; discuss the impact of different specimens on test performance; and provide practical advice to help clinicians and researchers new to the field select a test that best suits their needs.