To provide a detailed assessment of micrometastases of colorectal cancer by anatomical mapping of regional lymph nodes (LNs), we analyzed 237 LNs from 11 patients with colorectal cancer by reverse transcription-PCR (RT-PCR) using carcinoembryonic antigen and cytokeratin 20 as genetic markers. All dissected LNs were mapped anatomically and subjected to detection assays for micrometastases. Immunohistochemical analysis was also performed using anti-pancytokeratin antibody AE1/AE3 to confirm the existence of occult cancer cells. By histological analysis, 20 of 237 LNs contained metastatic cells, and they were all positive by both immunohistochemistry and RT-PCR. Of the 217 histologically negative LNs, 14 (6.5%) harbored micrometastases by immunohistochemistry, and 57 (26.2%) were positive for at least one of the two genetic markers. Lymphatic mappings of all patients showed that micrometastases were distributed not only at the pericolic LNs but often at distant LNS: Clinical follow-up study showed that two patients developed recurrence within 1 year after surgery, and both of them had RT-PCR-positive micrometastases in not less than 70% of LNs examined. Moreover, both patients had frequent micrometastases at distant LNs, i.e., those around the root or along the inferior mesenteric artery, when compared with patients with no recurrence. Our findings suggest that genetic diagnosis using the RT-PCR method may be clinically useful along with conventional pathological diagnosis, especially when micrometastases spread to distant LNS: