Recognition-Activated Primer-Mediated Exponential Rolling Circle Amplification for Signal Probe Production and Ultrasensitive Visual Detection of Ochratoxin A with Nucleic Acid Lateral Flow Strips

核酸 检出限 适体 化学 底漆(化妆品) 滚动圆复制 信号(编程语言) 核酸检测 赭曲霉毒素A 色谱法 生物系统 DNA DNA聚合酶 分子生物学 生物化学 真菌毒素 计算机科学 有机化学 生物 程序设计语言 食品科学
作者
Shi‐Yi Wang,Ziwen Zong,Jianguo Xu,Bangben Yao,Zhou Xu,Yao Li,Wei Chen
出处
期刊:Analytical Chemistry [American Chemical Society]
卷期号:95 (44): 16398-16406 被引量:10
标识
DOI:10.1021/acs.analchem.3c03995
摘要

We proposed a visual strategy for rapid and ultrasensitive detection of ochratoxin A (OTA) by integration of primer-mediated exponential rolling circle amplification (P-ERCA) with a designed nucleic acid lateral flow strip (LFS). The recognition component was preimmobilized in the tube by hybridization between the immobilized functionalized aptamer and complementary ssDNA. Recognition of OTA induces the release of complementary ssDNA from the tube, which will also act as the primer of the designed P-ERCA. Three nicking sites on the template P-ERCA could contribute to the production of enormous signal probes based on the simultaneous amplification-nicking model, which can be visually measured directly with the constructed nucleic acid LFS. Importantly, the nicked signal probe can also act as the trigger of the new-round RCA, achieving exponential growth of signal probes for measurement and signal enhancement. Taking advantage of the extraordinary amplification efficiency of P-ERCA and the simplicity of LFS, this P-ERCA-LFS method demonstrates ultrasensitive detection of OTA with a visual limit of detection as low as 100 fg/mL for qualitative screening and a limit of detection of 35 fg/mL for semiquantitative analysis. This designed strategy could also be utilized as a universal method for detection of other chemical analytes with the replacement of the aptamer for recognition, and the nucleic acid LFS unit could also be a useful protocol for direct ssDNA analysis.
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