title: Quantitative assessment of HER-2/neu protein concentration in breast cancer by enzyme-linked immunosorbent assay.
abstract: OBJECTIVE
  The HER-2/neu protein (p185) has become a promising target for antibody therapy in breast cancer. We tested the feasibility of a quantitative approach for HER-2/neu testing based on the analysis of tumor tissue extracts by an enzyme-linked immunosorbent assay (ELISA).
  
  
  METHODS
  Tumor tissue extracts of primary human breast cancers (n=124) were prepared using a triton-based buffer. HER-2/neu concentration was quantified by ELISA. Paraffin-embedded tissue sections of the same tumors were analyzed by immunohistochemical staining applying the monoclonal HER-2/neu antibody TAB 250 (n=124) and by chromogenic in situ hybridization (CISH) (n=73).
  
  
  RESULTS
  Concentrations of p185 in tissue extracts determined by ELISA varied from 1 to 927 ng per mg protein with a median of 25 ng/mg protein, whereas normal breast tissue showed values from 0.4 to 5.5 ng/mg with a median of 2.2 ng/mg (p<0.0001, Mann-Whitney U test). A significant correlation between p185 concentration and immunohistochemical staining was observed (p<0.0001, Kruskal-Wallis test). In addition, p185 concentration measured by ELISA was correlated with the degree of HER-2/neu gene amplification determined by CISH. HER-2/neu-amplified tumors had significantly higher p185 concentrations (median value 181 ng/mg protein) than non-amplified tumors (median value 20 ng/mg; p<0.0001, Mann-Whitney U test).
  
  
  CONCLUSIONS
  ELISA-based measurement of HER-2/neu protein concentration in breast cancer tissue extracts is feasible and provides quantitative results for p185 protein concentrations that correlate closely with HER-2/neu immunoscore and gene amplification.
authors.affiliation: Department of Obstetrics and Gynecology, University Hospital Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: V
authors.initials: V
authors.lastname: Müller
authors.forename: C
authors.initials: C
authors.lastname: Thomssen
authors.forename: C
authors.initials: C
authors.lastname: Karakas
authors.forename: I
authors.initials: I
authors.lastname: Eustermann
authors.forename: J
authors.initials: J
authors.lastname: Ramirez Porras
authors.forename: C
authors.initials: C
authors.lastname: Coith
authors.forename: J
authors.initials: J
authors.lastname: Berger
authors.forename: T
authors.initials: T
authors.lastname: Löning
authors.forename: F
authors.initials: F
authors.lastname: Jänicke
authors.forename: K
authors.initials: K
authors.lastname: Pantel
mesh_terms: D000911:Antibodies, Monoclonal; D001943:Breast Neoplasms / Q000378:metabolism*; D002875:Chromosomes / Q000648:ultrastructure; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis*; D013997:Time Factors
chemical_list: D000911:Antibodies, Monoclonal; D018719:Erb-b2 Receptor Tyrosine Kinases