Esophageal adenocarcinoma risk in Barrett's esophagus (BE) is increased 30- to 125-fold versus the general population. Among all BE patients, however, neoplastic progression occurs only once per 200 patient-years. Molecular biomarkers are therefore needed to risk-stratify patients for more efficient surveillance endoscopy and to improve the early detection of progression. We therefore performed a retrospective, multicenter, double-blinded validation study of eight BE progression prediction methylation biomarkers. Progression or nonprogression were determined at 2 years (tier 1) and 4 years (tier 2). Methylation was assayed in 145 nonprogressors and 50 progressors using real-time quantitative methylation-specific PCR. Progressors were significantly older than nonprogressors (70.6 versus 62.5 years; P < 0.001). We evaluated a linear combination of the eight markers, using coefficients from a multivariate logistic regression analysis. Areas under the ROC curve (AUC) were high in the 2-year, 4-year, and combined data models (0.843, 0.829, and 0.840; P < 0.001, <0.001, and <0.001, respectively). In addition, even after rigorous overfitting correction, the incremental AUCs contributed by panels based on the 8 markers plus age versus age alone were substantial (Delta-AUC = 0.152, 0.114, and 0.118, respectively) in all 3 models. A methylation biomarker-based panel to predict neoplastic progression in BE has potential clinical value in improving both the efficiency of surveillance endoscopy and the early detection of neoplasia.