A Novel Ultrasensitive Hybridization-Based ELISA Method for 2-Methoxyphosphorothiolate MicroRNAs and Its In vitro and In vivo Application
A Novel Ultrasensitive Hybridization-Based ELISA Method for 2-Methoxyphosphorothiolate MicroRNAs and Its In vitro and In vivo Application
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DOI：
10.1208/s12248-010-9214-0
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发表时间：
2010-12-01
期刊：
影响因子：
4.5
通讯作者：
Marcucci, Guido
中科院分区：
文献类型：
--
作者：
Chan, Kenneth K.;Liu, Zhongfa;Marcucci, Guido
MicroRNAs （miRNAs）是内源性的非编码小rna，与靶mrna结合并调节其表达。最近的证据表明，mirna参与人类恶性肿瘤。有研究表明，异常下调或上调的mirna可能分别被合成的mirna或拮抗剂mirna取代，并恢复正常的细胞功能。由于治疗发展需要分析支持，我们首次开发并验证了一种超灵敏和选择性的测定方法，用于定量小鼠血浆和细胞裂解液中合成的2'-甲氧基磷硫代酸- mirna。该方法基于杂交-连接荧光酶联免疫吸附测定，为三个合成mirna提供了10-1,000,000 pM的线性动态范围，包括单个和混合物。日内和日间变异系数分别为
MicroRNAs (miRNAs) are endogenous, small non-coding RNAs that bind to target mRNAs and regulate their expression. Recent evidence has indicated the involvement of miRNAs in human malignancies. It has been suggested that aberrantly down-regulated or up-regulated miRNAs may be replaced with synthetic miRNAs or antagomiRNAs, respectively, and restore normal cell functions. As therapeutic development requires analytical support, we developed and validated an ultrasensitive and selective assay for quantification of synthetic 2'-methoxyphosphorothiolate-miRNA in mouse plasma and cell lysate for the first time. The method is based on a hybridization-ligation fluorescence enzyme-linked immunosorbent assay and has provided a linear dynamic range of 10-1,000,000 pM for three synthetic miRNAs both singly and in a mixture. The intra- and inter-day coefficients of variation were