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Real‐time reverse transcription‐PCR and fluorescence in‐situ hybridization are complementary to understand the mechanisms involved in HER‐2/ neu overexpression in human breast carcinomas
Author(s) -
Vanden Bempt I,
Vanhentenrijk V,
Drijkoningen M,
Wlodarska I,
Vandenberghe P,
De WolfPeeters C
Publication year - 2005
Publication title -
histopathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.626
H-Index - 124
eISSN - 1365-2559
pISSN - 0309-0167
DOI - 10.1111/j.1365-2559.2005.02112.x
Subject(s) - fluorescence in situ hybridization , in situ hybridization , in situ , reverse transcription polymerase chain reaction , immunohistochemistry , reverse transcriptase , fluorescence , real time polymerase chain reaction , pathology , biology , microbiology and biotechnology , polymerase chain reaction , medicine , messenger rna , chemistry , gene , genetics , physics , organic chemistry , quantum mechanics , chromosome
Aims : To evaluate the HER‐2/ neu status at the mRNA and DNA level of breast carcinomas and to compare it with HER‐2/ neu receptor overexpression by immunohistochemistry (IHC). Methods and results : In 32 invasive breast carcinomas, frozen tissue was available for real‐time detection of HER‐2/ neu mRNA levels by reverse transcription‐polymerase chain reaction (RT‐PCR). Corresponding paraffin sections were examined by IHC and fluorescence in‐situ hybridization (FISH). Thereby, different IHC and FISH procedures were compared. Using microwave epitope retrieval, all 32 cases scored 3+ on IHC, whereas only 28 out of 32 cases scored IHC 3+ using water bath epitope retrieval. All of these 28 cases showed increased levels of HER‐2/ neu mRNA. Dual‐colour FISH analysis showed corresponding gene amplification in all 28 cases, with two cases showing a peculiar amplification pattern. In the remaining four cases, scoring IHC 2+ using water bath epitope retrieval, mRNA levels were not elevated. Three cases did not have gene amplification and one case showed low‐level HER‐2/ neu gene amplification. All four carcinomas showed chromosome 17 polysomy. Conclusions : Real‐time RT‐PCR is accurate in selecting breast carcinoma cases scoring 3+ by IHC with high‐level gene amplification. Results obtained by dual‐colour FISH suggest that mechanisms leading to HER‐2/ neu receptor overexpression may be different between carcinomas scoring 2+ and 3+ on IHC, with polysomy 17 found in the former and gene amplification in the latter.