核酸探针杂交及酶联显色方法定量检测微小RNA-21  

Quantitative detection of microRNA-21 by nucleic acid probe hybridization and enzyme-linked chromogenic assay

作　　者：于天晓 许红[1] 韩彦洁[1] 张瑶[1] 王亚芳[1] 张静蕊 李美芳[1] 腰利云 YU Tianxiao;XU Hong;HAN Yanjie;ZHANG Yao;WANG Yafang;ZHANG Jingrui;LI Meifang;YAO Liyun(Department of Blood Transfusion, the Fourth Hospital of Shijiazhuang/Shijiazhuang Maternity Hospital,Shijiazhuang 050000, China)

机构地区：[1]石家庄市第四医院/石家庄市妇产医院输血科

出　　处：《现代医学》2019年第7期805-807,共3页Modern Medical Journal

基　　金：河北省重点研发计划自筹项目(172777192)

摘　　要：目的:建立一种基于核酸探针杂交及酶联显色的高灵敏方法,实现对微小RNA-21(miRNA-21)的定性和定量检测。方法:将生物素标记的miRNA-21捕获探针通过生物素-亲和素系统固定在96孔板上,当靶miRNA-21出现在反应体系中时,能够与固定在96孔板上的捕获探针结合。利用辣根过氧化物酶标记的DNA检测探针的催化和信号放大作用,催化底物3,3’,5,5’-四甲基联苯胺,使反应体系出现颜色变化,根据颜色变化对miRNA-21的浓度进行定性和定量分析。结果:通过考察方法的灵敏度、精密度、回收率可得该方法最低检测限可达0. 81 pmol·L-1,相对标准偏差均在7. 5%以下,回收率在78. 9%~119. 2%之间,结论:本研究建立了一种高通量显色分析方法,该方法能够高灵敏检测miRNA-21,且具有良好的灵敏度、重复性和回收率。Objective: To established a high sensitivity method based on nucleic acid probe hybridization and enzyme-linked chromogenic for qualitative and quantitative detection of microRNA-21( miRNA-21). Methods:Biotin labelled miRNA-21 capture probe was immobilized on 96-well plate by biotin-streptavidin system. The capture probe could effectively bind to the target miRNA-21 which appeared in the reaction system. DNA detection probe labeled by horseradish peroxidase which had catalytic activity and signal amplification could catalytic substrate 3,3 ’,5,5 ’-tetramethylbenzidine to form a colorimetric reaction. Qualitative and quantitative analysis of the miRNA-21 concentration were performed based on the color change of reaction system. Result: By investigating the sensitivity,precision and recovery of the method,the lowest detection limit was 0. 81 pmol · L-1,relative standard deviationwere less than 7. 5%,and the recoveries were between 78. 9%-119. 2%. Conclusion: In this study,a sensitive and high throughput chromogenic method for detection of miRNA-21 is established. It has good sensitivity,repeatability and recovery.

关 键 词：微小RNA-21 检测方法 显色分析 

分 类 号：R-33[医药卫生]