A fully integrated, automated and rapid detection system for KRAS mutations

KRAS mutations are detected in tumors of various organs, and they are also markers of resistance for epidermal growth factor receptor (EGER) tyrosine kinase inhibitors and monoclonal antibodies against the EGFR. Thus, the accurate and rapid detection of KRAS mutations is crucial, not only for screening, but also for the prediction of the efficacy of molecular-targeted therapy. The aim of the present study was to establish a novel automated detection system for KRAS mutations. One hundred and thirty-six lung adenocarcinoma patients were genotyped for KRAS mutations with both the conventional direct sequence (DS) method and with the newly developed quenching probe (QP) method that obtains data automatically within 60 min. The detection limit of the QP method using a control plasmid containing the KRAS mutation was 50 copies, and 10% mutant plasmid was detected in the mixture of wild-type and mutants. The results obtained by the QP and DS methods were identical in all but two of the 136 cases. The two differentially identified samples, which consisted of substantially fewer lung cancer cells, were positive according to the QP method but negative as determined by DS for KRAS mutations. These findings characterize the QP method as an accurate and rapid detection system for KRAS mutations.