Colorectal cancer is the second-leading cause of cancer death. New noninvasive options for screening capable of diagnosing cancer at an early stage are needed to improve compliance and reduce mortality. This study was designed to provide an estimate of the sensitivity and specificity of a multitarget assay panel (MTAP) of stool DNA changes. Eighty patients with advanced colorectal neoplasia and 212 control subjects provided stool samples before colonoscopy. Patients with hereditary colorectal cancer syndromes were excluded. The MTAP included 21 specific mutations in the adenomatous polyposis coli (APC), p53, and K-ras genes, a microsatellite instability marker (BAT-26), and a marker of abnormal apoptosis (DNA Integrity Assay). All samples were analyzed in the clinical laboratory at EXACT Sciences. Multitarget assay panel detected 33 of 52 patients (63.5%, 95% confidence interval [CI], 49.0%-76.4%) with invasive colorectal cancer, including 26 of 36 (72.2%) with node-negative disease (American Joint Committee on Cancer [AJCC] stage I/II) and 7 of 16 (43.7%) with advanced disease (AJCC stage III/IV). Sixteen of 28 patients (57.1%; 95% CI, 37.2%-75.5%) with advanced adenomas (lesions containing high-grade dysplasia, villous adenomas, or tubular adenomas > 1 cm in size) were detected, including 6 of 7 (85.7%) with high-grade dysplasia and 10 of 21 (47.6%) with other advanced adenomas. Specificity was 96.2% (95% CI, 92.7%-98.4%) in patients with either no colorectal lesions or diminutive polyps. Multitarget assay panel has better sensitivity than that reported with use of Hemoccult(R) II in fecal occult blood testing, with similar specificity. Sensitivity appeared to be equally high for patients with node-negative and advanced disease, as well as for advanced adenomas. This study contained a disproportionately high number of distal cancers and, as such, may not be representative of results in proximal lesions. Although a prospective study in an average-risk population is needed to validate these findings, MTAP may offer an important noninvasive option for population-based screening.