Advanced Cell Diagnostics, Inc., Hayward, CA 94545, USA.
J Mol Diagn. 2012 Jan;14(1):22-9. doi: 10.1016/j.jmoldx.2011.08.002.
In situ analysis of biomarkers is highly desirable in molecular pathology because it allows the examination of biomarker status within the histopathological context of clinical specimens. Immunohistochemistry and DNA in situ hybridization (ISH) are widely used in clinical settings to assess protein and DNA biomarkers, respectively, but clinical use of in situ RNA analysis is rare. This disparity is especially notable when considering the abundance of RNA biomarkers discovered through whole-genome expression profiling. This is largely due to the high degree of technical complexity and insufficient sensitivity and specificity of current RNA ISH techniques. Here, we describe RNAscope, a novel RNA ISH technology with a unique probe design strategy that allows simultaneous signal amplification and background suppression to achieve single-molecule visualization while preserving tissue morphology. RNAscope is compatible with routine formalin-fixed, paraffin-embedded tissue specimens and can use either conventional chromogenic dyes for bright-field microscopy or fluorescent dyes for multiplex analysis. Unlike grind-and-bind RNA analysis methods such as real-time RT-PCR, RNAscope brings the benefits of in situ analysis to RNA biomarkers and may enable rapid development of RNA ISH-based molecular diagnostic assays.
在分子病理学中，对生物标志物进行原位分析是非常理想的，因为它可以在临床标本的组织病理学背景下检查生物标志物的状态。免疫组织化学和 DNA 原位杂交 (ISH) 广泛应用于临床，分别用于评估蛋白质和 DNA 生物标志物，但原位 RNA 分析的临床应用很少。当考虑到通过全基因组表达谱发现的大量 RNA 生物标志物时，这种差异尤其明显。这主要是由于当前 RNA ISH 技术的技术复杂性高、灵敏度和特异性不足。在这里，我们描述了 RNAscope，这是一种新型的 RNA ISH 技术，具有独特的探针设计策略，允许同时进行信号放大和背景抑制，以实现单分子可视化，同时保持组织形态。RNAscope 与常规的福尔马林固定、石蜡包埋组织标本兼容，并且可以使用传统的显色染料进行明场显微镜检查，也可以使用荧光染料进行多重分析。与实时 RT-PCR 等研磨结合的 RNA 分析方法不同，RNAscope 将原位分析的优势引入到 RNA 生物标志物中，并可能促进基于 RNA ISH 的分子诊断检测方法的快速发展。
