Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States.
Anal Chem. 2011 Sep 15;83(18):7086-93. doi: 10.1021/ac201307d. Epub 2011 Aug 17.
We describe an electrokinetic concentration-enhanced aptamer affinity probe electrophoresis assay to achieve highly sensitive and quantitative detection of protein targets in a microfluidic device. The key weaknesses of aptamer as a binding agent (weak binding strength/fast target dissociation) were counteracted by continuous injection of fresh sample while band-broadening phenomena were minimized due to self-focusing effects. With 30 min of continuous signal enhancement, we can detect 4.4 pM human immunoglobulin E (IgE) and 9 pM human immunodeficiency virus 1 reverse transcriptase (HIV-1 RT), which are among the lowest limits of detection (LOD) reported. IgE was detected in serum sample with a LOD of 39 pM due to nonspecific interactions between aptamers and serum proteins. The method presented in this paper also has broad applicability to improve sensitivities of various other mobility shift assays.
我们描述了一种电泳分析方法,通过电动增强亲和探针来实现微流控装置中对蛋白质靶标的高灵敏度和定量检测。通过持续注入新鲜样本,克服了适体作为结合剂的关键弱点(结合强度弱/目标快速解离),同时由于自聚焦效应,最小化了带展宽现象。通过 30 分钟的连续信号增强,可以检测到 4.4 pM 的人免疫球蛋白 E(IgE)和 9 pM 的人类免疫缺陷病毒 1 逆转录酶(HIV-1 RT),这是报告的最低检测限(LOD)之一。由于适体与血清蛋白之间的非特异性相互作用,IgE 在血清样本中的检测限为 39 pM。本文提出的方法也广泛适用于提高各种其他迁移率变动分析的灵敏度。