title: An alternative and reliable real-time quantitative PCR method to determine HER2/neu amplification in breast cancer.
abstract: The overexpression of HER-2/neu is an independent prognostic factor of clinical outcome of breast cancer, therefore determination of HER-2/neu status is now an integral part of the clinicopathologic workup. The ways of measuring the copy number of the HER-2/neu gene in tumor cells comprise in situ hybridization techniques and real-time polymerase chain reaction (PCR). Quantitative real-time PCR is a relatively new technique for assessing HER-2/neu gene amplification with high sensitivity. However, the HER-2/neu Quantification Kit developed by Roche designed for a LightCycler 1.5 platform had been withdrawn from the commercial market; therefore, we were encouraged to design an alternative LightCycler-based method that offers the desired level of reliability. One hundred breast cancer cases with known HER-2/neu status have been examined with the original Roche developed HER-2/neu Quantification kit and the custom real-time PCR assay. The newly developed, custom PCR showed sensitivity of 91.43%, specificity of 90.63%, and accuracy of 90.91% taking fluorescence in situ hybridization results as the end point. We have described a novel real-time PCR technique for the relative quantification of the HER2/neu gene on a LightCycler 1.5 platform. We have determined that our method is eligible and ideal for the supplement of regular fluorescence in situ hybridization reactions, concerning its high sensitivity and reliability.
authors.affiliation: Department of Pathology, Medical and Health Science Centre, University of Debrecen, Debrecen, Hungary.
authors.forename: Kristof
authors.initials: K
authors.lastname: Egervari
authors.forename: Judit
authors.initials: J
authors.lastname: Toth
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Nemes
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Szollosi
mesh_terms: D001483:Base Sequence; D001943:Breast Neoplasms / Q000175:diagnosis* / Q000235:genetics; D005755:Gastrins / Q000235:genetics; D005784:Gene Amplification; D018628:Gene Dosage; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011379:Prognosis; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D012680:Sensitivity and Specificity
chemical_list: D005755:Gastrins; D018719:Erb-b2 Receptor Tyrosine Kinases