Detection of c-Ki-ras mutations in faecal samples from sporadic colorectal cancer patients

Abstract

Colonic exfoliated epithelial cells in faecal material provide a source of human DNA which has been analysed for the presence of the tumour marker ras, in order to detect early tumour cells. The stool samples were subjected to a preliminary sample preparation step followed by centrifugation. DNA was extracted from both the centrifugation pellet and supernatant fractions, as well as from endoscopy washings, using a conventional phenol chloroform extraction method and was then purified on glass milk or spin columns. The purified DNA was amplified using mitochondrial primers and analysed for ras mutations using a non-radioactive, allele specific mismatch method. Corresponding tumour DNA was analysed for mutations using the same method. The results show that approximately 50% of the faecal samples analysed exhibited the presence of ras mutations which were also observed in the corresponding tumours. A double mutation was detected in one supernatant. Our findings represent an important stage in the development of a diagnostic test for the early detection of colorectal cancer.