Fluorescence Polarization Combined Capillary Electrophoresis Immunoassay for the Sensitive Detection of Genomic DNA Methylation

CORC > 生态环境研究中心 > 中国科学院生态环境研究中心 > 环境化学与生态毒理学国家重点实验室

	Fluorescence Polarization Combined Capillary Electrophoresis Immunoassay for the Sensitive Detection of Genomic DNA Methylation
	Wang, Xiaoli ; Song, Yuling ; Song, Maoyong ; Wang, Zhixin ; Li, Tao ; Wang, Hailin
刊名	ANALYTICAL CHEMISTRY
	2009-10-01
卷号	81 期号:19 页码:7885-7891
中文摘要	Genomic DNA hypomethylation is epigenetically associated with aberrant gene expression and chromosome instability. Here we describe a method for rapid and sensitive detection of genomic DNA methylation without the need for bisulfite conversion, enzymatic digestion, or PCR amplification. The methylated DNA is first specifically recognized by an anti-5-methylcytosine IgG(1) antibody and noncovalently labeled by a monovalent, fluorescently labeled, Fc-specific anti-IgG(1) Fab fragment (secondary antibody). The formed immunocomplex of methylated DNA can be efficiently focused and separated from the DNA unbound secondary antibody by capillary electrophoresis (CE). The free fluorescent dye comigrates with the immuno-complex. However, by taking advantage of online laser-induced fluorescence polarization detection (LIFP), the target immuno-complex can be distinguished and accurately measured from the overlapped dye without further separation. The developed method is highly sensitive with a LOD of 0.3 nM and is highly specific for the detection of methylated DNA. Moreover, the CE-LIFP immunoassay is rapid (1.2 min for one analysis) and only consumes less than 0.1 ng of genomic DNA. The method was validated by examining human cell lines treated by methyltransferase inhibitor 5-aza-2'-deoxycytidine at low doses (10 nM-10 mu M). Because of its sensitivity and speed, our method will be applicable for rapid epigenetic evaluation and for the study of tumorigenesis and chemical-cell interactions.
WOS记录号	WOS:000270361100009
内容类型	期刊论文
源URL	[http://ir.rcees.ac.cn/handle/311016/21504]
专题	生态环境研究中心_环境化学与生态毒理学国家重点实验室
推荐引用方式 GB/T 7714	Wang, Xiaoli,Song, Yuling,Song, Maoyong,et al. Fluorescence Polarization Combined Capillary Electrophoresis Immunoassay for the Sensitive Detection of Genomic DNA Methylation[J]. ANALYTICAL CHEMISTRY,2009,81(19):7885-7891.
APA	Wang, Xiaoli,Song, Yuling,Song, Maoyong,Wang, Zhixin,Li, Tao,&Wang, Hailin.(2009).Fluorescence Polarization Combined Capillary Electrophoresis Immunoassay for the Sensitive Detection of Genomic DNA Methylation.ANALYTICAL CHEMISTRY,81(19),7885-7891.
MLA	Wang, Xiaoli,et al."Fluorescence Polarization Combined Capillary Electrophoresis Immunoassay for the Sensitive Detection of Genomic DNA Methylation".ANALYTICAL CHEMISTRY 81.19(2009):7885-7891.