A site-specific DNA methylation biosensor for both visual and magnetic determination based on lateral flow assay
A site-specific DNA methylation biosensor for both visual and magnetic determination based on lateral flow assay
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一种基于侧流分析的位点特异性 DNA 甲基化生物传感器，用于视觉和磁性测定
作者：
Zhang Jiaxing;Liu Xiaonan;Zhang Sinong;Cai Yu;Ma Kang;Hua Kai;Cui Yali
由DNA甲基化异常驱动的肿瘤发生已经突显了开发一种便携式、快速和灵敏的策略来利用特定的癌症预后基因进行准确的甲基化检测的必要性，以迎合精确医学的普及。在本研究中，建立了一种基于侧向流动分析的DNA甲基化视觉和磁性位点特异性生物传感器。通过将地高辛和生物素标记的引物引入到聚合酶链式反应中，该生物传感器中的金磁性纳米颗粒(GMNPs)可以识别和捕获扩增产物。作为一种信号探针，GMNPs的光学性质允许用肉眼解释扩增产物，避免了任何复杂的设备和繁琐的操作。此外，由于GMNP的磁性，在临床上可以用磁强计来解释和记录信号。特制的引物组的引入使得在大量未甲基化的变异体作为强干扰背景的情况下准确区分0.1%的甲基化变异体，反之亦然。对于视觉和磁性分析，低至0.01pg的变种都可以观察到明显的信号。作为一个重要的抑癌基因，miR-34a的启动子甲基化状态不仅可以用细胞株准确测定，也可以用临床标本准确测定，显示了该生物传感器在肿瘤诊断和预后方面的巨大潜力。
Tumorigenesis driven by abnormal DNA methylation has highlighted the need to develop a portable, rapid and sensitive strategy for accurate methylation detection with a specific cancer-prognostic gene, which caters to the popularization of precision medicine. In this study, a site-specific biosensor for both visual and magnetic DNA methylation determination has been established based on lateral flow assay. By introducing digoxin- and biotin-labeled primers into PCR, the amplicons can be recognized and captured by gold magnetic nanoparticles (GMNPs) in this biosensor. Working as a signal probe, the optical property of GMNPs allows the amplicons to be interpreted with naked eyes avoiding any complex equipment and cumbersome operation after PCR. Moreover, by virtue of the magnetic property of GMNP, the signal can be explained and recorded by a magnetometer in clinical practice. The introduction of tailor-made primer sets makes it possible to accurately distinguish 0.1% methylated variants in the presence of numerous unmethylated variants as strong interferential background and vice versa at target cytosine–guanine dinucleotide. A distinct signal can be observed with as low as 0.01 pg variants for both visual and magnetic analyses. As a significant tumor suppressor gene, the promoter methylation status of miR-34a is accurately determined with not only cell lines but also with clinical samples, which demonstrates the great potential of this biosensor for cancer diagnosis and prognosis.