期刊
ANALYTICAL CHEMISTRY
卷 88, 期 22, 页码 11108-11114出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.6b03163
关键词
-
资金
- NSFC [21273117, 21375063, 21335004]
- Program for Outstanding Innovation Research Team of Universities in Jiangsu Province
- Priority Academic Program Development of Jiangsu Higher Education Institutions
- Jiangsu Government
Effective detection of DNA methyltransferase (DNMT) activity is significant for cancer research. Herein, we developed a sensitive electroanalytical method to detect human DNA (cytosine-5) methyltransferase 1 (DNMT1) from crude lysates of cancer cells. In this assay, capture DNA having a preferred DNMT1 methylation site was immobilized on a gold electrode and then hybridized with gold nanoparticle (Au NP)-DNA complexes. The modified electrodes were equilibrated with the lysate and then incubated with methylation-sensitive restriction enzyme. If the lysate was negative for DNMT1 activity, the Au NP DNA complexes would be cut by the restriction enzyme and released from the electrode. Conversely, restriction enzyme cleavage would be blocked by the fully methylated duplexes, and the Au NP DNA complexes would remain on the electrode. Electroactive Ru(NH3)(6)(3+) was used as the signal reporter, because of its electrostatic attraction to DNA, resulting in an electrochemical signal. Since the electrochemical signal reflects the amount of Ru(III) redox and the amount of Ru(III) redox is correlated with the activity of DNMT1, the activity of DNMT1 is proportional to the electrochemical signal. The signal could be amplified by the numerous DNAs on the Au NPs and further amplified by Ru(III) redox recycling. With this method, a detection limit down to 0.3 U/mL for pure DNMT1 and 8 MCF-7 cells was achieved. DNMT1 activities of different cell lines were also successfully evaluated.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据