Purpose To evaluate the feasibility and repeatability of various metabolically active tumor volume (MATV) quantification methods in fluorine 18 fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) in a multicenter setting and propose the optimal MATV method together with the minimal threshold for future response evaluation studies. Materials and Methods The study was approved by the institutional review board of all four participating centers, and patients provided written informed consent. Thirty-four patients with advanced gastrointestinal malignancies underwent two FDG PET/CT examinations within 1 week. MATVs were defined semiautomatically with 27 variations of tumor delineation methods with different reference values. Feasibility was determined as the percentage of successful tumor segmentations per MATV method. Repeatability was determined with intraclass correlation coefficients, Bland-Altman plots, and limits of agreement (LOAs) of the percentage difference between the test and repeat test measurements. In addition, LOA variability per center was investigated. Results In total, 136 lesions were identified. Feasibility of tumor segmentation ranged from 54% to 100% (74-136 of 136 lesions); repeatability was evaluated for 19 MATV methods with feasibility of greater than 95%. The median MATV derived with 50% threshold of mean standardized uptake value (SUV) of a sphere of 12-mm diameter with highest local intensity (SUVhp), which may not include the voxel with highest SUV corrected for local background, was 5.7 and 6.1 mL for test and retest scans, respectively, with a relative LOA of 36.1%. Comparable repeatability was found between centers. A difference in uptake time between scan 1 and 2 of 15 minutes or longer had a minor negative influence on repeatability. Conclusion MATV measured with 50% of SUVhp corrected for local background is recommended in multicenter FDG PET/CT studies on the basis of a high feasibility (96%) and repeatability (LOA of 36.1%). © RSNA, 2014 Online supplemental material is available for this article.