Real-time quantitative PCR of microdissected paraffin-embedded breast carcinoma: A alternative method for HER-2/neu analysis

Lise Mette Gjerdrum, Boe Sandahl Sorensen, Eigil Kjeldsen, Flemming Brandt Sorensen, Ebba Nexo, Stephen Hamilton-Dutoit

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    Abstrakt

    We studied the feasibility of using real-time quantitative PCR to determine HER-2 DNA amplification and mRNA expression in microdissected formalin-fixed, paraffin-embedded breast tumors and compared this with standard immunohistochemistry (MC) and fluorescent in situ hybridization (FISH) methods. Study cases (27 carcinoma and 3 ductal breast carcinoma in situ (DCIS) cases) showed varying Her-2 expression as determined by IHC (HercepTest). In carcinomas, there was a good correlation between HER-2 DNA amplification and strong HER-2 protein expression detected by FISH and IHC, respectively. A single DCIS case was amplified in FISH, but not in IHC. Both HER-2 gene amplification and expression could be quantified in microdissected paraffin-embedded tumors using real-time PCR, DNA and RNA being successfully detected in 146 of 150 (97%) and 141 of 150 (94%) samples, respectively. PCR analysis for HER-2 DNA amplification using the LightCycler HER2/neu DNA Quantification kit (Roche Molecular Biochemicals, Mannheim, Germany) correlated fairly well with IHC and FISH. All IHC HER-2 3+ tumors were amplified according to the kit, as was the FISH-amplified DCIS case. DNA-PCR identified five additional tumors as being amplified. Interestingly, all these scored 2+ with the HercepTest, but were negative using FISH. We believe that real-time quantitative PCR analysis of HER-2 DNA amplification following microdissection represents a useful supplementary or perhaps even an alternative technique for establishing HER-2 status in paraffin-embedded tumors.

    OriginalsprogEngelsk
    Sider (fra-til)42-51
    Antal sider10
    TidsskriftJournal of Molecular Diagnostics
    Vol/bind6
    Udgave nummer1
    DOI
    StatusUdgivet - feb. 2004

    Fingeraftryk Udforsk hvilke forskningsemner 'Real-time quantitative PCR of microdissected paraffin-embedded breast carcinoma: A alternative method for HER-2/neu analysis' indeholder.

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