Postoperative survival of colorectal cancer patients is often delineated by metastases spreading to the liver. Current clinical diagnostic procedures are unable to discover micrometastases in this organ. Our aim was to develop a diagnostic tool for detecting micrometastases that are present at the time of surgery. Therefore, a PCR-RFLP assay was set up tracking point mutations of the K-ras oncogene at codons 12 and 13, based on mismatch primers and restriction enzymes BstXI and XcmI. The detection limit of this assay was one mutant in one million wild-type cells. One hundred forty-two patients with colorectal carcinoma were screened for these mutations in tissue samples from their tumor, proximally adjacent mucosa, and liver. Of these, 67 patients (46%) were positive for a K-ras mutation, of which 58 had codon 12 and 9 had codon 13 mutations. No patient without a K-ras-positive tumor showed a mutation in mucosa, but 11 patients with a K-ras-positive tumor (11 of 58; 19%) were found to bear a K-ras mutation in their mucosa, and in 21 patients (21 of 64; 33%), a K-ras mutation was detected in liver tissue. Sequencing of all mutated samples revealed a 92% confirmation of PCR-RFLP results. In summary, the assay is a useful tool for detecting K-ras codon 12 and 13 mutations and allows early proof of molecular determinants of liver metastases. Such knowledge will improve the staging of colorectal cancer patients and could beneficially influence their prognosis if followed by an effective therapy.