Chromogenic in situ hybridization -: A practical alternative for fluorescence in situ hybridization to detect HER-2/neu oncogene amplification in archival breast cancer samples
Chromogenic in situ hybridization -: A practical alternative for fluorescence in situ hybridization to detect HER-2/neu oncogene amplification in archival breast cancer samples
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DOI：
10.1016/s0002-9440(10)64785-2
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发表时间：
2000-11-01
影响因子：
6
通讯作者：
Isola, J
中科院分区：
文献类型：
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作者：
Tanner, M;Gancberg, D;Isola, J
HER-2/neu癌基因扩增的测定已成为选择乳腺癌患者进行曲妥珠单抗（赫赛汀）治疗的必要条件。荧光原位杂交（FISH）是目前检测HER 2/neu扩增的金标准方法，但在常规组织病理学实验室中并不十分实用。我们评估了一种新的修改原位杂交，显色原位杂交（CISH），这使得检测HER-2/neu基因拷贝与传统的过氧化物酶反应。预处理（通过在微波炉中加热和使用酶消化）福尔马林固定石蜡包埋的肿瘤组织切片，并与地高辛标记的DNA探针杂交。用抗地高辛荧光素、抗荧光素过氧化物酶和二氨基联苯胺检测探针。通过CISH可视化的基因拷贝可以在苏木精染色的组织切片中用x40物镜容易地区分。HER-2/neu扩增通常表现为大的过氧化物酶阳性核内基因拷贝簇。CISH和FISH（根据Vysis，由冷冻粉碎的肿瘤样本制成）在一系列157例乳腺癌中相关性良好（kappa系数为0.81）。结果表明，CISH和CB-11免疫组化检测均为阴性，而FISH检测结果均为阴性。结论：CISH是一种检测HER-2/neu扩增的有效方法，尤其是对免疫组化结果的确认。
Determination of HER-2/neu oncogene amplification has become necessary for selection of breast cancer patients for trastuzumab (Herceptin) therapy. Fluorescence in situ hybridization (FISH) is currently regarded as a gold standard method for detecting HER2/neu amplification, but it is not very practical for routine histopathological laboratories. We evaluated a new modification of in situ hybridization, the chromogenic ill situ hybridization (CISH), which enables detection of HER-2/neu gene copies with conventional peroxidase reaction. Archival formalin-fixed paraffin-embedded tumor tissue sections were pretreated (by heating In a microwave oven and using enzyme digestion) and hybridized with a digoxigenin-labeled DNA probe. The probe was detected with antidigoxigenin fluorescein, anti-fluorescein peroxidase, and diaminobenzidine. Gene copies visualized by CISH could be easily distinguished with a x40 objective in hematoxylin-stained tissue sections. HER-2/neu amplification typically appeared as large peroxidase-positive intranuclear gene copy clusters. CISH and FISH (according to Vysis, made from frozen pulverized tumor samples) correlated well in a series of 157 breast cancers (kappa coefficient, 0.81). The few different classifications were mostly because of low-level amplifications by FISH that were negative by CISH and immunohistochemistry with monoclonal antibody CB-11, We conclude that CISH, using conventional bright-field microscopy in evaluation, is a useful alternative for determination of HER-2/neu amplification in paraffin-embedded tumor samples, especially for confirming the immunohistochemical staining results.