RNAscope for in situ detection of transcriptionally active human papillomavirus in head and neck squamous cell carcinoma.
RNAscope for in situ detection of transcriptionally active human papillomavirus in head and neck squamous cell carcinoma.
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DOI：
10.3791/51426
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发表时间：
2014-03-11
期刊：
影响因子：
--
通讯作者：
Ma XJ
中科院分区：
文献类型：
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作者：
Wang H;Wang MX;Su N;Wang LC;Wu X;Bui S;Nielsen A;Vo HT;Nguyen N;Luo Y;Ma XJ
致癌HPV检测的“金标准”是肿瘤组织中转录活性高风险HPV的证明。然而，通过定量逆转录聚合酶链反应（qRT-PCR）检测E6/E7 mRNA需要RNA提取，这破坏了对形态学相关性至关重要的肿瘤组织背景，并且难以在常规临床实践中采用。我们最近开发的RNA原位杂交技术，RNAscope，允许直接可视化的RNA在福尔马林固定，石蜡包埋（FFPE）组织与单分子灵敏度和单细胞分辨率，这使得高灵敏度和特异性原位分析的任何RNA生物标志物在常规临床标本。RNAscope HPV检测试剂盒设计用于使用基因型特异性探针池检测7种高危HPV基因型（HPV 16、18、31、33、35、52和58）的E6/E7 mRNA。相对于目前通过qRT-PCR检测E6/E7 mRNA的“金标准”方法，该方法已表现出优异的灵敏度和特异性。通过RNAscope确定的HPV状态是口咽癌患者临床结局的强预后。
The 'gold standard' for oncogenic HPV detection is the demonstration of transcriptionally active high-risk HPV in tumor tissue. However, detection of E6/E7 mRNA by quantitative reverse transcription polymerase chain reaction (qRT-PCR) requires RNA extraction which destroys the tumor tissue context critical for morphological correlation and has been difficult to be adopted in routine clinical practice. Our recently developed RNA in situ hybridization technology, RNAscope, permits direct visualization of RNA in formalin-fixed, paraffin-embedded (FFPE) tissue with single molecule sensitivity and single cell resolution, which enables highly sensitive and specific in situ analysis of any RNA biomarker in routine clinical specimens. The RNAscope HPV assay was designed to detect the E6/E7 mRNA of seven high-risk HPV genotypes (HPV16, 18, 31, 33, 35, 52, and 58) using a pool of genotype-specific probes. It has demonstrated excellent sensitivity and specificity against the current 'gold standard' method of detecting E6/E7 mRNA by qRT-PCR. HPV status determined by RNAscope is strongly prognostic of clinical outcome in oropharyngeal cancer patients.