Backgrounds: Delay in the diagnosis of tuberculosis (TB) leads to poor response to treatment and the disease transmission to susceptible individuals. The Xpert MTB/RIF assay efficiently detects Mycobacterium tuberculosis (MTB). The present study aimed to compare acid-fast bacilli (AFB) microscopy, culture, and Xpert MTB/RIF assay in the diagnosis of pulmonary and extrapulmonary tuberculosis cases. Materials & Methods: This retrospective study was conducted in the Department of Microbiology, Government Medical College, Srinagar, India over 18 months from February 2019 to July 2020. Samples were processed and evaluated using AFB microscopy, culture, and Xpert MTB/RIF assay.Findings: Among the 1862 samples evaluated, 224 samples were found to be positive using AFB microscopy, culture, and Xpert MTB/RIF assay. The overall sensitivity and specificity of the Xpert MTB/RIF assay in diagnosing pulmonary TB cases was 98.23 and 97.69%, respectively. Among the smear-negative extrapulmonary samples, 52 (5.75%) and 86 (9.6%) samples were positive in culture and the Xpert MTB/RIF assay, respectively. The maximum recovery of MTB by Xpert MTB/RIF assay was from tissue biopsy specimens. Rifampicin resistance was observed in six samples. Conclusion: Both culture and Xpert MTB/RIF methods were sensitive in detecting smearpositive samples. Although both techniques missed some smear-negative pulmonary and extrapulmonary TB cases, the Xpert MTB/RIF assay enhanced the detection rate of MTB compared to culture. The Xpert MTB/RIF assay enabled the accurate diagnosis of tuberculosis cases with a rapid turnaround time; therefore, it could assist clinicians to start timely therapeutic interventions for these patients.