title: Detection of mutations by fill-in ligation reaction with enzyme-linked immunosorbent assay for rapid medical diagnosis.
abstract: Several approaches for parallel genotyping have been developed with increasingly available information on DNA variation. However, these methods require either complex laboratory procedures or expensive instrumentation. None of these procedures is readily performed in local clinical laboratories. In this study, we developed a flexible genotyping method involving fill-in ligation reaction with enzyme-linked immunosorbent assay successfully applied to detect important single-nucleotide polymorphisms (SNPs) for EGFR c.2573T > G (L858R), EGFR c.2582T > A (L861Q), and EGFR c.2155G > T (G719C). This assay exhibited excellent specificity, with a sensitivity as low as 0.5%. Eight out of 62 clinical samples were identified as heterozygotes for the SNP site of L858R, whereas only two samples were identified as heterozygotes by direct sequencing. The developed method enabled accurate identification of SNP in a simple and cost-effective manner adapted to routine analysis.
authors.affiliation: a Medical College of Hubei University of Arts and Science , XiangYang , China.
authors.forename: Yi-Tong
authors.initials: YT
authors.lastname: Tang
authors.forename: Na
authors.initials: N
authors.lastname: Xiao
authors.forename: Zhi-Shan
authors.initials: ZS
authors.lastname: Li
authors.forename: Jiu-Ming
authors.initials: JM
authors.lastname: Zou
authors.forename: Rui
authors.initials: R
authors.lastname: Cao
authors.forename: Xue-Hong
authors.initials: XH
authors.lastname: Zhao
authors.forename: Jin-Hui
authors.initials: JH
authors.lastname: Shao
mesh_terms: D001483:Base Sequence; D004252:DNA Mutational Analysis / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000235:genetics; D005091:Exons / Q000235:genetics; D000077156:Gefitinib; D060005:Genotyping Techniques; D006801:Humans; D008175:Lung Neoplasms / Q000097:blood / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D010957:Plasmids / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D011799:Quinazolines / Q000494:pharmacology / Q000627:therapeutic use; D013997:Time Factors; D016896:Treatment Outcome
chemical_list: D011799:Quinazolines; C512478:EGFR protein, human; D066246:ErbB Receptors; D000077156:Gefitinib
keywords: EGFR; SNP; fill-in ligation reaction; genotyping; mutations