Reactive oxygen species (ROS) are at the centre of many physiological and pathological processes. NOX1, a ROS-producing NADPH oxidase, is highly expressed in the colon but its function in colonic physiology or pathology is still poorly understood. It has been suggested to play a role in host defence, but also in cell growth and possibly malignant transformation. In this study we characterized NOX1 expression in human colon samples derived from healthy control subjects and patients with colon cancer or inflammatory bowel disease (IBD). NOX1 mRNA expression was assessed by dot-blot hybridization, real-time PCR and in situ hybridization, using samples derived from surgical specimens from patients undergoing colon resection. In normal tissues, NOX1 expression was low in the ileum, intermediate in the right colon, and high in the left colon (p = 0.0056 right vs. left colon). NOX1 mRNA levels were not influenced by factors linked to colon tumourigenesis, such as age or sex. Moreover, there was no statistical difference in NOX1 expression between samples derived from adenomas, well differentiated or poorly differentiated colon adenocarcinomas. At a cellular level, NOX1 was highly expressed in colon epithelial cells, both within the crypts and on the luminal surface. In addition, a population of lymphocytes, particularly in the appendix, showed NOX1 expression. Lymphocytes in lesions of Crohn's disease and ulcerative colitis were also strongly positive for NOX1. In conclusion, NOX1 is an enzyme that is constitutively expressed in colon epithelium and is not associated with tumourigenesis. Its distribution in crypts and on the luminal surface, as well as its left-to-right gradient in the colon, suggests a role in host defence function. In addition to the known epithelial localization, we define lymphocytes as a novel site of NOX1 expression, where it may potentially be involved in the pathogenesis of inflammatory bowel diseases.