Guidelines for the use of nucleic acid amplification (NAA) tests for the diagnosis of tuberculosis (TB) were published in 1996 and updated in 2000. Since then, NAA testing has become a routine procedure in many settings because NAA tests can reliably detect Mycobacterium tuberculosis bacteria in specimens 1 or more weeks earlier than culture. Earlier laboratory confirmation of TB can lead to earlier treatment initiation, improved patient outcomes, increased opportunities to interrupt transmission, and more effective public health interventions. Because of the increasing use of NAA tests and the potential impact on patient care and public health, in June 2008, CDC and the Association of Public Health Laboratories (APHL) convened a panel of clinicians, laboratorians, and TB control officials to assess existing guidelines and make recommendations for using NAA tests for laboratory confirmation of TB. On the basis of the panel's report and consultations with the Advisory Council for the Elimination of TB (ACET),* CDC recommends that NAA testing be performed on at least one respiratory specimen from each patient with signs and symptoms of pulmonary TB for whom a diagnosis of TB is being considered but has not yet been established, and for whom the test result would alter case management or TB control activities, such as contact investigations. These guidelines update the previously published guidelines.