Objective: Oral Squamous Cell Carcinoma (SCCA) diagnosis in at-risk populations has proven resistant to reduction in morbidity and mortality. To change this pattern, we study enhanced appreciation of at-risk sites and accelerated identification of asymptomatic lesions by Tolonium chloride. Method: Tolonium chloride stain was tested by sequential visual exam, then stain for lesions arising in previously treated SCCA surgical and radiation fields in a 3 year trial of diagnostic efficacy in 13 clinical centers. Visual exam, the Gold standard, was followed by Tolonium chloride stain, read by a second, blinded examiner. Protocol demanded repetition of positive exams +10-20 days, unless urgent biopsy was dictated by first exam. We tested effectiveness and exam success rates by nonparametric statistics. Oral Pathologists, blinded to clinical exam results, provided histologic diagnoses, which constituted definitive diagnoses. Results: From 396 lesions, 96 biopsies were taken (12.1% total patients with positive lesions), of which 30 (31.3%) were diagnosed as SCCA/carcinoma-in-situ, and the remainder inflammation (31.3%), keratosis (26.6%), dysplasia (21.9%) and ulcer (2.1%). Of SCCA, 12 (40.0%) were considered to be clinically suspicious. Nearly all SCCA (25/26, 96.2%) and a majority of erythroplakic lesions (23/36, 63.9%) were considered suspicious. Progressively lower percentages of lesions diagnosed as nodular leukoplakia (6/13, 46.2%), lichen planus (8/34, 23.5%), benign leukoplakia (11/78, 14.1%), inflammation (4/38, 10.5%), and late radiation change (3/41, 7.3%) were directed for biopsy. Tolonium chloride sensitivity for SCCA=0.98 (p=0.0002), Visual exam sensitivity =0.40, Positive Predictive Value =36%, without Kappa difference. Conclusion: Tolonium chloride enhances identification of abnormal mucosal sites by experienced clinicians, may promote biopsy, assist in biopsy site selection, accelerate early diagnosis and identify unsuspicious dysplastic lesions. This multi-center study indicated sensitivity of Tolonium chloride in detection of all lesions while maintaining excellent SCCA sensitivity as compared to clinical exam alone. Support: Zila, Inc.