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.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
This novel molecular approach can be performed in <5 h facilitating intraoperative use for real-time surgical resection.
Harden
Susan V
SV
Department of Otolaryngology-Head and Neck Surgery, The Johns Hopkins University School of Medicine, Baltimore, Maryland, 21205, USA.
Thomas
David C
DC
Benoit
Nicole
N
Minhas
Khalid
K
Westra
William H
WH
Califano
Joseph A
JA
Koch
Wayne
W
Sidransky
David
D
D002294:Carcinoma, Squamous Cell / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004252:DNA Mutational Analysis / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D016158:Genes, p53; D005821:Genetic Techniques; D006258:Head and Neck Neoplasms / Q000235:genetics; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics; D008198:Lymph Nodes / Q000473:pathology; D008207:Lymphatic Metastasis; D009693:Nucleic Acid Hybridization; D017354:Point Mutation; D013997:Time Factors
D017931:DNA Primers; D004247:DNA