Mucosal biopsy criteria has limited validity in terms of discrimination between ulcerative colitis (UC) and Crohn's disease (CD). The aim of this study was to set up quantitative immunohistochemical criteria, with a special focus on inflammatory cell distribution within individual specimens and throughout the large bowel. Quantitative evaluation was performed for the density of CD8+, CD45RO+, neutrophil elastase+, CD68+ and mast cell tryptase+ cells in affected and unaffected mucosa taken from 41 patients with UC and 61 patients with CD. Each slide was examined at the highest and lowest density fields, which were further divided into the upper and deeper half of mucosa. Multiple logistic regression analysis using 51 features as independent variables constructed a predictive equation finding the probability of UC (PUC), and the diagnostic categories were subsequently defined based on a receiver-operating characteristic curve. The analysis disclosed five significant features suggesting UC; these implied intense infiltration of CD8+ and mast cell tryptase+ cells, diffuse infiltration of neutrophil elastase+ and CD68+ cells, and continuous infiltration of CD45RO+ cells. The criteria consisted of three diagnostic categories, 'suggestive of UC (PUC > or = 0.7)', 'indeterminate (0.3 < PUC < 0.7)', and 'suggestive of CD (PUC < or = 0.3)'; the criteria had values for sensitivity and specificity exceeding 95%. The immunohistochemical criteria distinguishing UC from CD may help to confirm the diagnosis in patients with ambiguous endoscopic and histological diagnosis.