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Graphene oxide-quenching-based fluorescence in situ hybridization (G-FISH) to detect RNA in tissue: Simple and fast tissue RNA diagnostics.
基于氧化石墨烯猝灭的荧光原位杂交 (G-FISH) 检测组织中的 RNA: 简单快速的组织 RNA 诊断。
Alzheimer's disease Formalin-fixed paraffin-embedded (FFPE) tissue Glioblastoma multiforme tumor Graphene oxide-quenching-based fluorescence in situ hybridization (G-FISH) Tissue RNA diagnostics
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摘要

FISH-based RNA detection in paraffin-embedded tissue can be challenging, with complicated procedures producing uncertain results and poor image quality. Here, we developed a robust RNA detection method based on graphene oxide (GO) quenching and recovery of fluorescence in situ hybridization (G-FISH) in formalin-fixed paraffin-embedded (FFPE) tissues. Using a fluorophore-labeled peptide nucleic acid (PNA) attached to GO, the endogenous long noncoding RNA BC1, the constitutive protein β-actin mRNA, and miR-124a and miR-21 could be detected in the cytoplasm of a normal mouse brain, primary cultured hippocampal neurons, an Alzheimer's disease model mouse brain, and glioblastoma multiforme tumor tissues, respectively. Coding and non-coding RNAs, either long or short, could be detected in deparaffinized FFPE or frozen tissues, as well as in clear lipid-exchanged anatomically rigid imaging/immunostaining-compatible tissue hydrogel (CLARITY)-transparent brain tissues. The fluorescence recovered by G-FISH correlated highly with the amount of miR-21, as measured by quantitative real time RT-PCR. We propose G-FISH as a simple, fast, inexpensive, and sensitive method for RNA detection, with a very low background, which could be applied to a variety of research or diagnostic purposes.

译文

石蜡包埋组织中基于 FISH 的 RNA 检测可能具有挑战性,复杂的程序会产生不确定的结果和较差的图像质量。在此,我们开发了一种基于氧化石墨烯 (GO) 猝灭和福尔马林固定石蜡包埋 (FFPE) 组织中荧光原位杂交 (G-FISH) 恢复的稳健的 RNA 检测方法。使用附着在 GO 上的荧光团标记的肽核酸 (PNA),内源性长链非编码 RNA BC1,组成蛋白 β-肌动蛋白 mRNA, 在正常小鼠大脑、原代培养的海马神经元、阿尔茨海默病模型小鼠大脑和多形性胶质母细胞瘤组织的细胞质中可以检测到 miR-124a 和 miR-21,分别。编码和非编码 rna,无论是长的还是短的,都可以在非咖啡因的 FFPE 或冷冻组织中检测到, 以及在透明脂质交换的解剖学刚性成像/免疫染色相容的组织水凝胶 (CLARITY) 透明脑组织中。通过定量实时 G-FISH 测量,miR-21 恢复的荧光与 RT-PCR 量高度相关。我们提出 G-FISH 是一种简单、快速、廉价和敏感的 RNA 检测方法,其背景非常低,可应用于各种研究或诊断目的。

Fluorescence in situ hybridization

肿瘤 基础研究 实验技术
概述  :  

荧光原位杂交技术,简称FISH,是利用荧光标记的特异核酸探针与细胞内相应的靶DNA或RNA分子杂交,通过在荧光显微镜或共聚焦激光扫描仪下观察荧光信号,来确定与特异探针杂交后被染色的细胞或细胞器的形态和分布,或者是结合了荧光探针的DNA区域或RNA分子在染色体或其他细胞器中的定位。该技术能够在组织、细胞等水平上对DNA或RNA进行定性、定位以及定量分析。原理FISH检测技术是一种重要的非放射性原位杂交技术,它的基本原理是:如果被检测的染色体或DNA纤维切片上的靶DNA与所用的核酸探针是同源互补

Fluorescence   英  [flʊə'res(ə)ns; flɔː-]   美  [flʊə'rɛsns]

       n. 荧光;荧光性

同根词   adj.fluorescent 荧光的;萤光的;发亮的

       This approach uses fluorescence instead of magnetism to image samples.  该方法利用的是荧光而非影像样本的磁性。

 

in   英  [ɪn]   美  [ɪn]

       prep. 在……内,在……里;进……里;在……期间;在……以后;穿着;在……状态中;在……方面;包含在……中;

               adv. 进入;在里头;到达;当选;上涨,涨潮;在屋里;在车站;收到;递交,送到;

               adj. 在里面的;时髦的;(用球)界内的

               n. 执政者;门路

       He was a big man, dressed in a suit and tie.  他是个高大的男人,穿着西服、打着领带。


situ   英  [sɪtju:]   美  [ˈsaɪtu]

       n. 原地

       Back to square one! Nothing stood in the distance vision in situ!  回到原点!站在原地憧憬一无所有的远方!

 

hybridization   英  [,haɪbrɪdaɪ'zeɪʃən]   美  [,haɪbrɪdɪ'zeʃən]

       n. 杂交;配种;[化学] 杂化;杂种培殖

同根词   adj.hybrid 混合的;杂种的

       Detection and analysis of the hybridization graph.  杂交图谱的检测和分析。

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