The diagnosis of infection following shoulder arthroplasty is notoriously difficult. The prevalence of prosthetic shoulder infection after arthroplasty ranges from 3.9 – 15.4% and the most common infective organism is Cutibacterium acnes. Current preoperative diagnostic tests fail to provide a reliable means of diagnosis including WBC, ESR, CRP and joint aspiration. Fluoroscopic-guided percutaneous synovial biopsy (PSB) has previously been reported in the context of a pilot study and demonstrated promising results. The purpose of this study was to determine the diagnostic accuracy of percutaneous synovial biopsy compared with open culture results (gold standard). This was a multicenter prospective cohort study involving four sites and 98 patients who underwent revision shoulder arthroplasty. The cohort was 60% female with a mean age was 65 years (range 36-83 years). Enrollment occurred between June 2014 and November 2021. Pre-operative fluoroscopy-guided synovial biopsies were carried out by musculoskeletal radiologists prior to revision surgery. A minimum of five synovial capsular tissue biopsies were obtained from five separate regions in the shoulder. Revision shoulder arthroplasty was performed by fellowship-trained shoulder surgeons. Intraoperative tissue samples were taken from five regions of the joint capsule during revision surgery. Of 98 patients who underwent revision surgery, 71 patients underwent both the synovial biopsy and open biopsy at time of revision surgery. Nineteen percent had positive infection based on PSB, and 22% had confirmed culture positive infections based on intra-operative tissue sampling. The diagnostic accuracy of PSB compared with open biopsy results were as follows: sensitivity 0.37 (95%CI 0.13-0.61), specificity 0.81 (95%CI 0.7-0.91), positive predictive value 0.37 (95%CI 0.13 – 0.61), negative predictive value 0.81 (95%CI 0.70-0.91), positive likelihood ratio 1.98 and negative likelihood ratio 0.77. A patient with a positive pre-operative PSB undergoing revision surgery had an 37% probability of having true positive infection. A patient with a negative pre-operative PSB has an 81% chance of being infection-free. PSB appears to be of value mainly in ruling out the presence of peri-prosthetic infection. However, poor likelihood ratios suggest that other ancillary tests are required in the pre-operative workup of the potentially infected patient.