Harper Cancer Research Institute, University of Notre Dame, 1234 Notre Dame Avenue, Harper Hall A212, Notre Dame, IN 46556, USA.
J Oncol. 2012;2012:903581. doi: 10.1155/2012/903581. Epub 2012 May 13.
The noncoding RNA designated as microRNA (miRNA) is a large group of small single-stranded regulatory RNA and has generated wide-spread interest in human disease studies. To facilitate delineating the role of microRNAs in cancer pathology, we sought to explore the feasibility of detecting microRNA expression in formalin-fixed paraffin-embedded (FFPE) tissues. Using FFPE materials, we have compared fluorescent in situ hybridization (FISH) procedures to detect miR-146a with (a) different synthetic probes: regular custom DNA oligonucleotides versus locked nucleic acid (LNA) incorporated DNA oligonucleotides; (b) different reporters for the probes: biotin versus digoxigenin (DIG); (c) different visualization: traditional versus tyramide signal amplification (TSA) system; (d) different blocking reagents for endogenous peroxidase. Finally, we performed miR-146a FISH on a commercially available oral cancer tissue microarray, which contains 40 cases of oral squamous cell carcinoma (OSCC) and 10 cases of normal epithelia from the human oral cavity. A sample FISH protocol for detecting miR-146a is provided. In summary, we have established reliable in situ hybridization procedures for detecting the expression of microRNA in FFPE oral cancer tissues. This method is an important tool for studies on the involvement of microRNA in oral cancer pathology and may have potential prognostic or diagnostic value.
非编码 RNA 被指定为 microRNA（miRNA），是一大组小的单链调节 RNA，在人类疾病研究中引起了广泛关注。为了促进阐明 microRNA 在癌症病理学中的作用，我们试图探索在福尔马林固定石蜡包埋（FFPE）组织中检测 microRNA 表达的可行性。使用 FFPE 材料，我们比较了荧光原位杂交（FISH）程序，以检测 miR-146a 与：（a）不同的合成探针：常规定制 DNA 寡核苷酸与锁核酸（LNA）掺入的 DNA 寡核苷酸；（b）探针的不同报告器：生物素与地高辛（DIG）；（c）不同的可视化：传统与酪胺信号放大（TSA）系统；（d）内源性过氧化物酶的不同阻断试剂。最后，我们对商业上可用的口腔癌组织微阵列进行了 miR-146a 的 FISH 分析，该阵列包含 40 例口腔鳞状细胞癌（OSCC）和 10 例来自人类口腔的正常上皮。提供了检测 miR-146a 的 FISH 方案示例。总之，我们已经建立了可靠的原位杂交程序，用于检测 FFPE 口腔癌组织中 microRNA 的表达。该方法是研究 microRNA 在口腔癌病理学中的作用的重要工具，可能具有潜在的预后或诊断价值。
