title: Validation of a differential PCR and an ELISA procedure in studying HER-2/neu status in breast cancer.
abstract: HER-2/neu oncogene status and total cellular p185HER-2 content were simultaneously analyzed in 415 invasive breast-cancer specimens by differential PCR and ELISA respectively. Mathematical analysis of the data led us to establish a cut-off value of 1.7 for the ratio between the intensity of the HER-2/neu gene band and the reference gene band, to consider the HER-2/neu gene amplified, and of 260 fmol/mg protein, to consider p185HER-2 over-expressed. Of the 415 tumors studied, 15% showed a diverse degree of HER-2/neu gene amplification. Of these tumors, 87% showed over-expression of the p185HER-2. Of the remaining 352 specimens that did not display HER-2/neu gene amplification, 97% showed no p185HER-2 over-expression (p < 0.0001). In 40 selected samples with a p185HER-2 level lower than 260 fmol/mg protein, the degree of p185HER-2 phosphorylation was very low or undetectable. Conversely, 38 of 46 selected tumors with a p185HER-2 level higher than 260 fmol/mg protein exhibited a considerable degree of p185HER-2 phosphorylation (p < 0.0001). Our data suggest that: (i) differential PCR and ELISA, which are relatively simple procedures, give similar information on HER-2/neu status in breast cancer; and (ii) given the large series analyzed, the cutoff values established can be considered as safe values for determining whether, in a given tumor, the HER-2/neu oncogene is amplified or p185HER-2 is over-expressed.
authors.affiliation: Department of Cellular and Molecular Endocrinology, Faculty of Medicine, University of Las Palmas de Gran Canaria, Spain.
authors.forename: P F
authors.initials: PF
authors.lastname: Valerón
authors.forename: R
authors.initials: R
authors.lastname: Chirino
authors.forename: L
authors.initials: L
authors.lastname: Fernandez
authors.forename: S
authors.initials: S
authors.lastname: Torres
authors.forename: D
authors.initials: D
authors.lastname: Navarro
authors.forename: J
authors.initials: J
authors.lastname: Aguiar
authors.forename: J J
authors.initials: JJ
authors.lastname: Cabrera
authors.forename: B N
authors.initials: BN
authors.lastname: Diaz-Chico
authors.forename: J C
authors.initials: JC
authors.lastname: Diaz-Chico
mesh_terms: D001483:Base Sequence; D001943:Breast Neoplasms / Q000378:metabolism*; D015342:DNA Probes; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis*; D014407:Tumor Cells, Cultured
chemical_list: D015342:DNA Probes; D018719:Erb-b2 Receptor Tyrosine Kinases