Detection of BRAF mutations in colorectal tumours and peritoneal washings using a mismatch ligation assay.
OBJECTIVE
To detect cells bearing BRAF mutations in colorectal tumour samples and peritoneal washings, using a mismatch ligation assay (MLA).


METHODS
DNA from 46 colorectal tumours was studied. Part of exon 15 of the BRAF gene was amplified using the polymerase chain reaction, and T-->A mutations at codon 600 were detected using MLA. When a mutation was detected, the same mutation was sought in peritoneal washings from that patient.


RESULTS
BRAF mutations were detected in five of the 45 analysable tumour samples. In four cases, this result was confirmed by sequencing analysis. More tumours with BRAF mutations were Dukes' stage C or D rather than A or B (p < 0.02). Dilution experiments revealed that one mutant cell could be detected in 1000 normal cells. Cells with the same BRAF mutation were present in the peritoneal washing taken at the start of the operation in four of the five patients.


CONCLUSIONS
MLA is a suitable technique for the detection of BRAF mutations, and allows the detection of small numbers of isolated tumour cells shed from the primary tumour.
Busby
K
K
University of Warwick, Coventry CV4 AL, UK.
Morris
A
A
D000368:Aged; D001202:Ascitic Fluid / Q000473:pathology; D001483:Base Sequence; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000473:pathology; D004273:DNA, Neoplasm / Q000235:genetics; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009154:Mutation / Q000235:genetics; D009363:Neoplasm Proteins / Q000235:genetics*; D009367:Neoplasm Staging; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D010533:Peritoneal Lavage / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*
D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf