OBJECTIVESTo compare two different methods to evaluate rectal epithelial cell proliferation as a biomarker of risk of developing colon cancer.METHODSSamples of normal rectal mucosa from 26 patients at increased risk for colorectal cancer (22 patients with adenoma, three with adenocarcinoma of the large bowel, and one with longstanding ulcerative colitis) were examined by means of in vitro labeling with tritiated thymidine and flow cytometry.RESULTSWe found a significant correlation between thymidine-labeling index and the percentage of cells in S-phase, measured by flow cytometry both in formalin-fixed, paraffin-embedded specimens and in frozen specimens (respectively, r = 0.7647, p < 0.001, and r = 0.4503, p < 0.01). However, using flow cytometry, the percentage of cells in S-phase was significantly higher than the thymidine-labeling index in both fixed-embedded and frozen specimens (p < 0.01). Proliferative parameters were not higher in patients with colon carcinoma, and were not related to the degree of dysplasia, the number of adenomas, or familial occurrence of colorectal cancer. Two specimens taken from normal rectal mucosa of two patients with adenomas showed aneuploidy. No aneuploidy was found in normal rectal specimens of patients with adenocarcinoma.CONCLUSIONSThese results show that the calculation of cells in S-phase with in vitro tritiated thymidine labeling or by flow cytometry produces different results. However, the significant correlation between corresponding parameters obtained with these techniques support the use of either method as "intermediate biomarkers" of colorectal cancer risk and prognosis.