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Background  The clinical benefit of determining the status of HER-2/ neu  amplification in breast cancer patients is well accepted. Although immunohistochemistry (IHC) is the most frequently used method to assess the over-expression of HER-2 protein, fluorescent  in-situ  hybridization (FISH) is recognized as the "gold standard" for the determining of HER-2/ neu  status. The greatest discordance between the two methods occurs among breast tumors that receive an indeterminate IHC score of 2+. More recently, a real-time polymerase chain reaction (PCR) assay using the LightCycler ®  has been developed for quantifying HER-2/ neu  gene amplification. In this study, we evaluated the sensitivity and specificity of a commercially available LightCycler assay as it compares to FISH. To determine whether this assay provides an accurate alternative for the determination of HER-2/ neu  status, we focused primarily on tumors that were deemed indeterminate or borderline status by IHC.    Methods  Thirty-nine breast tumors receiving an IHC score of 2+ were evaluated by both FISH and LightCycler ®  technologies in order to determine whether quantitative real-time PCR provides an accurate alternative for the determination of HER-2/ neu  status.    Results  We found a high concordance (92%) between FISH and real-time PCR results. We also observed that 10% of these tumors were positive for gene amplification by both FISH and real-time PCR.    Conclusion  The data show that the results obtained for the gene amplification of HER-2/ neu  by real-time PCR on the LightCycler ®  instrument is comparable to results obtained by FISH. These results therefore suggest that real-time PCR analysis, using the LightCycler ® , is a viable alternative to FISH for reassessing breast tumors which receive an IHC score of 2+, and that a combined IHC and real-time PCR approach for the determination of HER-2 status in breast cancer patients may be an effective and efficient strategy.

Real-time PCR complements immunohistochemistry in the determination of HER-2/neu status in breast cancer