The reliability of the BDProbeTec ET assay, a semiautomated system for the direct detection of Mycobacterium tuberculosis complex in clinical specimens was evaluated by comparing results obtained by the system to those obtained by conventional culture methods and clinical data of the patients. Out of 1131 (735 respiratory and 396 nonrespiratory) specimens, 42 were smear- and 125 culture-positive. After resolution of discrepant results the overall sensitivity, specificity, and positive and negative predictive values for the BDProbeTec ET assay were 90.3, 96.9, 78.3, and 98.9%, respectively. For smear-positive specimens, all values were 100%, while the values for smear-negative specimens were 86.1, 96.9, 70.1, and 98.8%, respectively. All 28 nontuberculous mycobacteria were negative with the BDProbeTec ET system. The data demonstrate that the BDProbeTec ET assay is a highly sensitive and specific technique for the rapid (within 5 h) detection of M. tuberculosis complex in respiratory and nonrespiratory specimens.