Real-time gap ligase chain reaction: a rapid semiquantitative assay for detecting p53 mutation at low levels in surgical margins and lymph nodes from resected lung and head and neck tumors.

Abstract

We have developed a real-time semiquantitative gap ligase chain reaction for detecting p53 point mutations at low level in a background of excess of wild-type DNA. This method was validated by direct comparison to a previously validated but cumbersome phage plaque hybridization assay. Forty-one surgical margins and lymph nodes from 10 cases of head and neck squamous cell carcinoma and lung carcinoma were tested for p53 mutant clones. Both methods detected p53 mutants in margins from 8 of the 10 cases, whereas standard pathology detected cancer cells in only 3 cases. Positive margins included tissue samples with a tumor/normal DNA ratio of up to 1:1000. This novel molecular approach can be performed in <5 h facilitating intraoperative use for real-time surgical resection.