适体
肉眼
化学
脱氧核酶
分析物
组合化学
毒品检测
检出限
计算生物学
小分子
纳米技术
色谱法
生物化学
分子生物学
生物
材料科学
作者
Yingping Luo,Haixiang Yu,Obtin Alkhamis,Yingzhu Liu,Xinhui Lou,Boyang Yu,Yi Xiao
标识
DOI:10.1021/acs.analchem.9b00507
摘要
Colorimetric aptamer-based sensors offer a simple means of on-site or point-of-care analyte detection. However, these sensors are largely incapable of achieving naked-eye detection, because of the poor performance of the target-recognition and signal-reporting elements employed. To address this problem, we report a generalizable strategy for engineering novel multimodule split DNA constructs termed “CBSAzymes” that utilize a cooperative binding split aptamer (CBSA) as a highly target-responsive bioreceptor and a new, highly active split DNAzyme as an efficient signal reporter. CBSAzymes consist of two fragments that remain separate in the absence of target, but effectively assemble in the presence of the target to form a complex that catalyzes the oxidation of 2,2′-azino-bis(3-ethylbenzthiazoline)-6-sulfonic acid, developing a dark green color within 5 min. Such assay enables rapid, sensitive, and visual detection of small molecules, which has not been achieved with any previously reported split-aptamer-DNAzyme conjugates. In an initial demonstration, we generate a cocaine-binding CBSAzyme that enables naked-eye detection of cocaine at concentrations as low as 10 μM. Notably, CBSAzyme engineering is straightforward and generalizable. We demonstrate this by developing a methylenedioxypyrovalerone (MDPV)-binding CBSAzyme for visual detection of MDPV and 10 other synthetic cathinones at low micromolar concentrations, even in biological samples. Given that CBSAzyme-based assays are simple, label-free, rapid, robust, and instrument-free, we believe that such assays should be readily applicable for on-site visual detection of various important small molecules such as illicit drugs, medical biomarkers, and toxins in various sample matrices.
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