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Purpose To validate the AO Spine Subaxial Injury Classification System with participants of various experience levels, subspecialties, and geographic regions. Methods A live webinar was organized in 2020 for validation of the AO Spine Subaxial Injury Classification System. The validation consisted of 41 unique subaxial cervical spine injuries with associated computed tomography scans and key images. Intraobserver reproducibility and interobserver reliability of the AO Spine Subaxial Injury Classification System were calculated for injury morphology, injury subtype, and facet injury. The reliability and reproducibility of the classification system were categorized as slight (ƙ = 0–0.20), fair (ƙ = 0.21–0.40), moderate (ƙ = 0.41–0.60), substantial (ƙ = 0.61–0.80), or excellent (ƙ = > 0.80) as determined by the Landis and Koch classification. Results A total of 203 AO Spine members participated in the AO Spine Subaxial Injury Classification System validation. The percent of participants accurately classifying each injury was over 90% for fracture morphology and fracture subtype on both assessments. The interobserver reliability for fracture morphology was excellent (ƙ = 0.87), while fracture subtype (ƙ = 0.80) and facet injury were substantial (ƙ = 0.74). The intraobserver reproducibility for fracture morphology and subtype were excellent (ƙ = 0.85, 0.88, respectively), while reproducibility for facet injuries was substantial (ƙ = 0.76). Conclusion The AO Spine Subaxial Injury Classification System demonstrated excellent interobserver reliability and intraobserver reproducibility for fracture morphology, substantial reliability and reproducibility for facet injuries, and excellent reproducibility with substantial reliability for injury subtype.