Background Uniportal video-assisted thoracic surgery (UVATS) technique has been increasingly used for many thoracic diseases. Whether UVATS has equivalent or better perioperative outcomes for pulmonary sequestration (PS) patients remains controversial. Our study aimed to evaluate the feasibility of UVATS in anatomical lung resection for pulmonary sequestration.Methods A total of 24 patients with PS including fifteen males and nine females with the mean age of 40.54±14.49 (rang, 18-65) years old, who had received completely UVATS anatomical lung resection for PS in Nanjing Chest Hospital between January 2016 and December 2018 were retrospectively reviewed. Referring clinical data were retrieved from hospital records and analyzed.Results All 24 patients had been treated with the UAVTS approach successfully with no aberrant artery ruptured and no massive hemorrhage occurred, and no patients died during the perioperative period. Overall median surgery time was 102.21 ±28.00 mins (range, 55-150mins), the mean blood loss was 94.17±78.90 ml (range, 10-300ml), the mean days of chest tube maintained were 4.21±2.21 days (range,1-10days), and the mean postoperative hospitalization days was 5.79 ±2.35 days (range,2-11days). All patients were cured, without cough, fever, hemoptysis, and so on, occurring during the average follow-up of 17.42 ±2.00 months (range, 3-35months).Conclusions Our preliminary results revealed that anatomical lung resection by UVATS is a safe and feasible mini-invasive technique for PS patients, which might be associated with less postoperative pain, reduced paresthesia, better cosmetic results, and faster recovery.