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Purpose: This study was performed to evaluate the factors affecting the diagnostic accuracy and complications of CT-guided automated cutting needle biopsy with extrapleural locating (EPL) method in mediastinal masses. Materials and method: A retrospective study of 155 consecutive mediastinal biopsies with confirmed final diagnosis was undertaken. All patients were performed CT-guided automated cutting needle biopsy(ACNB) using an EPL method. Final diagnoses were based on a retrospective analysis of surgical outcomes, results of biopsy and clinical follow-up lasting at least 6 months. Benign and malignant biopsy results were compared with final diagnoses to determine the diagnostic accuracy of the method. A statistical analysis of factors related to lesion size, lesion location, approach to lesion, number of biopsy, method of histopathology and final diagnosis (benign or malignant) was performed to determine possible influences on diagnostic accuracy. A p value less than 0.05 was interpreted as statistically significant. All complications were evaluated and analyzed. Results: CT-guided ACNB using EPL method provided adequate samples in 146/155 cases in the first biopsy procedure, with a total sample rate of 94.2%. 9 cases were performed a second biopsy because of inadequate samples. There were 121 malignant and 34 benign lesions. Sensitivity, specificity, positive predictive value, negative predictive value and accuracy for a diagnosis of malignancy were 93.4%, 79.4%, 94.2%, 77.1%, respectively. Overall diagnostic accuracy was 90.3%. Factors affecting diagnostic accuracy were lesion size (lesions≤3cm 79.2%, lesions>3cm 96.1%; p<0.05), method of histopathology (hematoxylin-eosin staining 81.3%, immunohistochemistry 100%; p<0.05) and final diagnosis (benign 79.4%, malignant 93.4%, p<0.05). There were 13 complications in 66 cases using transpulmonary approaches (13/66, 19.7%). Pneumothorax was the most common complication (10/66, 15.2%). In two cases, they required placement of a chest drain. There were three cases of a very small amount of haemoptysis (3/66, 4.5%). There were no complications in 89 cases with direct mediastinal approaches. Conclusions: In CT-guided ACNB using EPL method of mediastinal masses, the lesion size, method of histopathology and final diagnosis affect diagnostic accuracy. The complications of CT-guided ACNB using EPL method of mediastinal masses are rare and mild. It is a safe and effective technique for obtaining tissues for pathologic evaluation.