Single-Molecule Kinetic Fingerprinting for the Ultrasensitive Detection of Small Molecules with Aptasensors

文献类型： 外文期刊

第一作者： Weng, Rui

作者： Weng, Rui;Qiu, Jing;Qian, Yongzhong;Lou, Shengting;Li, Lidan;Zhang, Yi;Su, Xin;Walter, Nils G.

作者机构：

期刊名称：ANALYTICAL CHEMISTRY （ 影响因子：6.986； 五年影响因子：6.755 ）

ISSN： 0003-2700

年卷期： 2019 年 91 卷 2 期

页码：

收录情况： SCI

摘要： Aptamers have emerged as promising molecular tools for small-molecule analyte sensing. However, the performance of such aptasensors is generally limited by leakage since it has been difficult to completely suppress signal in the absence of analyte, resulting in a compromise between sensitivity and specificity. Here, we describe a methodology for the ultrasensitive detection of analytes combining aptasensors with single-molecule kinetic fingerprinting. A short, fluorescently labeled DNA probe is utilized to detect the structural changes upon ligand binding to the designed hairpin-shaped aptasensor probe. The Poisson statistics of binding and dissociation events of the DNA probe to single surface-immobilized aptasensor molecules is monitored by total internal reflection fluorescence microscopy, permitting the high-accuracy discrimination of the ligand bound and ligand-free states, resulting in zero background. The programmable dynamics of the hairpin enables fine-tuning of the hybridization kinetics of the fluorescent probe, rendering the acquisition time sufficiently flexible to optimize discrimination. Remarkable detection limits are achieved for a diverse set of analytes when spiked into chicken meat extract: the nucleotide adenosine (0.3 pM), the insecticide acetamiprid (0.35 pM), and the dioxin-like toxin PCB-77 (0.72 pM), which is superior to recently reported aptasensors. Our generalizable method significantly improves the performance of aptasensors, with the potential to extend to other molecular biomarkers.

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