Abstract: Objective: To establish a method of calibration and performance verification of the GCMS-QP 2010 Ultra gas chromatography-mass spectrometer for urine organic acid detection after annual maintenance, so as to ensure that the performance of the detection system can continuously meet the detection needs. Methods: According to the requirements of the equipment manufacturer and the Calibration Specifications for Benchtop Gas Chromatography-Mass Spectrometer, the gas chromatography-mass spectrometer was calibrated after annual maintenance. According to the CNAS-CL02-A003 Guidance on the Application of Accreditation Criteria for the Medical Laboratory Quality and Competence in the Field of Clinical Chemistry, the analytical performance of the maintained gas chromatography-mass spectrometer was validated. Results: The calibration results of quality range, quality accuracy, resolution, signal-to-noise ratio and repeatability were all met the requirements. The detection results of internal quality control materials were under control, and the results of retained samples kept unchanged. Conclusion: The calibration and performance verification method of the gas chromatography-mass spectrometer for urine organic acid detection after annual maintenance is established successfully, which is of importance for providing accurate and reliable results of urine organic acid.