Gastrointestinal (GI) tumors continue to be major causes of cancer-related mortality, in part, reflecting metastases that escape detection by histopathology. Moreover, although approximately 10% of carcinomas arise from unknown locations, these tumors frequently originate in the GI tract. Guanylyl cyclase C (GC-C) is a receptor selectively expressed by intestinal epithelial cells whose persistent expression by colorectal carcinomas and ectopic expression by adenocarcinomas of the upper GI tract suggest its use as a marker for GI malignancies. Here, expression of GC-C protein, identified by immunohistochemistry, was examined in tissues and tumors arising from the human GI tract. Guanylyl cyclase C protein was expressed by epithelial cells from the duodenum to the rectum, but not by those in normal esophagus and stomach. Expression was retained in tubular adenomas, inflammatory bowel disease, premalignant lesions, and in primary and metastatic adenocarcinomas from the colon, including metastases to lymph nodes and liver. Moreover, GC-C was ectopically expressed in all cases of dysplasia and adenocarcinomas arising from intestinal metaplasia in esophagus and stomach. Thus, GC-C appears to be an immunohistochemical marker for identifying adenocarcinomas of unknown origin, metastases in patients undergoing staging for GI adenocarcinomas, and intestinal metaplasia, dysplasia, and tumors arising therein in the upper GI tract.