An alternative and reliable real-time quantitative PCR method to determine HER2/neu amplification in breast cancer.
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 However, the HER-2/neu Quantification Kit developed by Roche designed for a LightCycler 1.5 platform had been withdrawn from the commercial market; therefore, we were encouraged to design an alternative LightCycler-based method that offers the desired level of reliability.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.
Egervari
Kristof
K
Department of Pathology, Medical and Health Science Centre, University of Debrecen, Debrecen, Hungary.
Toth
Judit
J
Nemes
Zoltan
Z
Szollosi
Zoltan
Z
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
D005755:Gastrins; D018719:Erb-b2 Receptor Tyrosine Kinases