Positive margins and margin clearance are risk factors for recurrence in oral cavity squamous cell carcinoma (OCSCC), and these features are used to guide decisions regarding adjuvant radiation treatment. However, the prognostic value of intraoperative tumor bed vs resection specimen sampling is not well defined. To determine the prognostic implications of intraoperative margin assessment methods (tumor bed vs resection specimen sampling) with recurrence among patients who undergo surgical resection for OCSCC. This was a retrospective study of patients who had undergone surgical resection of OCSCC between January 1, 2000, and December 31, 2021, at a tertiary-level academic institution. Patients were grouped by margin assessment method (tumor bed [defect] or resection specimen sampling). Of 223 patients with OCSCC, 109 patients had localized tumors (pT1-T2, cN0), 154 had advanced tumors, and 40 were included in both cohorts. Disease recurrence after surgery was estimated by the cumulative incidence method and compared between cohorts using hazard ratios (HRs). Data analyses were performed from January 5, 2023, to April 30, 2023. Recurrence-free survival (RFS). The study population comprised 223 patients (mean [SD] age, 62.7 [12.0] years; 88 (39.5%) female and 200 [90.0%] White individuals) of whom 158 (70.9%) had defect-driven and 65 (29.1%) had specimen-driven margin sampling. Among the 109 patients with localized cancer, intraoperative positive margins were found in 5 of 67 (7.5%) vs 8 of 42 (19.0%) for defect- vs specimen-driven sampling, respectively. Final positive margins were 3.0% for defect- (2 of 67) and 2.4% for specimen-driven (1 of 42) margin assessment. Among the 154 patients with advanced cancer, intraoperative positive margins were found in 29 of 114 (25.4%) vs 13 of 40 (32.5%) for defect- and specimen-driven margins, respectively. Final positive margins were higher in the defect-driven group (9 of 114 [7.9%] vs 1 of 40 [2.5%]). When stratified by margin assessment method, the 3-year rates of local recurrence (9.7% vs 5.1%; HR, 1.37; 95% CI, 0.51-3.66), regional recurrence (11.0% vs 10.4%; HR, 0.85; 95% CI, 0.37-1.94), and distant recurrence (6.4% vs 5.0%; HR, 1.10; 95% CI, 0.36-3.35) were not different for defect- vs specimen-driven sampling cohorts, respectively. The 3-year rate of any recurrence was 18.9% in the defect- and 15.2% in the specimen-driven cohort (HR, 0.93; 95% CI, 0.48-1.81). There were no differences in cumulative incidence of disease recurrence when comparing defect- vs specimen-driven cases. The findings of this retrospective cohort study indicate that margin assessment methods using either defect- or specimen-driven sampling did not demonstrate a clear association with the risk of recurrence after OCSCC resection. Specimen-driven sampling may be associated with reduced surgical margin positivity rates, which often necessitate concurrent chemotherapy with adjuvant radiation therapy.