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Objective: Resistance against a wide variety of antibiotics is one of the prominent characteristics of Acinetobacter baumannii. The purpose of this study was to investigate the in vitro susceptibility testing of A. baumannii isolates to rifampin and the examination of the value of routine antibiogram with disk diffusion, E-test, and agar dilution methods on collected isolates from a tertiary hospital in north-west Iran. Materials and Methods: Susceptibility of 68 clinically isolated A. baumannii against rifampin using three in vitro methods was investigated. For the E-test method, the Pachon–Ibanez's and Saballs's study criteria were used. The Pachon–Ibamez criteria were used for agar dilution method. For disk diffusion, the standard Kirby–Bauer diffusion method was used. The area under curve (AUC) was used to determine the appropriate methods. The methods were interpreted using sensitivity, specificity, and negative and positive predictive values. Results: A. baumannii susceptibility to the rifampin according to the E-test was 41.2% (Panchon–Ibanez criteria) and 32.4% (Saballs's criteria). The susceptibility was 29.4% according to the agar dilution method for the Panchon–Ibanez criteria, 2.9% according to the agar dilution method for the Saballs's criteria and, and 1.5% according to the disk diffusion methods. The results of the E-test method according to Pachon–Ibanez's and Saballs's criteria in comparison with the result of the agar dilution method according to Pachon–Ibanez's criteria had the highest AUC. Conclusion: According to the susceptibility testing of rifampin against A. baumannii, the E-test method has a higher diagnostic value than the agar dilution and disk diffusion methods.