title: The combinatorial approach of laser-captured microdissection and reverse transcription quantitative polymerase chain reaction accurately determines HER2 status in breast cancer.
abstract: BACKGROUND
  HER2 expression in breast cancer correlates with increased metastatic potential, higher tumor recurrence rates and improved response to targeted therapies. Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) are two methods commonly used for the analysis of HER2 in the clinic. However, lack of standardization, technical variability in laboratory protocols and subjective interpretation are major problems associated with these testing procedures.
  
  
  METHODS
  Here we evaluated the applicability of reverse-transcription quantitative polymerase chain reaction (RT-qPCR) for HER2 testing in breast cancer. We tested thirty formaldehyde-fixed and paraffin-embedded tumor samples by RT-qPCR, FISH and IHC and analysed and compared the data from the three methods.
  
  
  RESULTS
  We found that laser-captured microdissection is essential for the accurate determination of HER2 expression by RT-qPCR. When isolating RNA from total tumor tissue we obtained a significant number of false negative results. However, when using RNA from purified cancer cells the RT-qPCR data were fully consistent with FISH and IHC. In addition we provide evidence that ductal carcinomas might be further classified by the differential expression of HER3 and HER4.
  
  
  CONCLUSIONS
  Laser-captured microdissection in combination with RT-qPCR is a precise and cost-effective diagnostic approach for HER2 testing in cancer. The PCR assay is simple, accurate and robust and can easily be implemented and standardized in clinical laboratories.
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Hofmann
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.|Pathology Laboratory Obrist and Brunhuber, Klostergasse 1, A-6511 Zams, Austria.
authors.forename: Rita
authors.initials: R
authors.lastname: Seeboeck
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Nico
authors.initials: N
authors.lastname: Jacobi
authors.affiliation: Pathology Laboratory Obrist and Brunhuber, Klostergasse 1, A-6511 Zams, Austria.
authors.forename: Peter
authors.initials: P
authors.lastname: Obrist
authors.affiliation: Pathology Laboratory Obrist and Brunhuber, Klostergasse 1, A-6511 Zams, Austria.
authors.forename: Samuel
authors.initials: S
authors.lastname: Huter
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Christian
authors.initials: C
authors.lastname: Klein
authors.affiliation: Research Program for Rational Drug Design in Dermatology and Rheumatology, Department of Dermatology, Paracelsus Medical University of Salzburg, Müllner Hauptstraße 48, A-5020 Salzburg, Austria.
authors.forename: Kamil
authors.initials: K
authors.lastname: Oender
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Christoph
authors.initials: C
authors.lastname: Wiesner
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Harald
authors.initials: H
authors.lastname: Hundsberger
authors.affiliation: Department Life Sciences, IMC University of Applied Sciences Krems, Piaristengasse 1, A-3500 Krems, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Eger
keywords: Breast cancer; HER2; Microdissection; Personalized medicine; Polymerase chain reaction