Use of the most rapid and reliable laboratory tests for mycobacterial detection, identification, and susceptibility testing is important for TB control. In 1993, CDC experts made recommendations regarding optimal methods of mycobacterial testing (i.e., stains for AFB, culture, identification, and susceptibility testing of M. tuberculosis) and turnaround times for reporting results. Various technical advances have enhanced the diagnostic capability of the laboratory and/or improved laboratory efficiency since then. The commercial NAA tests for direct detection of MTBC have the greatest potential to impact patient care. To assist physicians, CDC experts have published recommendations concerning use of the NAA tests for management of patients with suspected TB, with emphasis on the MTD assay, which is approved for both AFB smear-positive and smear-negative specimens. With regard to mycobacterial culture, totally automated, nonradiometric systems are commercially available. For mycobacterial identification, various molecular techniques have been developed, but at present, they are used predominantly in research or large reference laboratories. Molecular tests also have proved useful for better understanding the epidemiology of TB and investigating episodes of suspected laboratory cross-contamination. With regard to mycobacterial susceptibility testing, use of the new automated culture systems for testing MTBC is under evaluation, but only one such system has been approved for this purpose. In addition, laboratory guidelines for susceptibility testing of MTBC and certain NTM have recently been published by the NCCLS.