Enhancing Point-of-Care Diagnosis of African Swine Fever Virus (ASFV) DNA with the CRISPR-Cas12a-Assisted Triplex Amplified Assay

文献类型： 外文期刊

第一作者： Zhu, Dan

作者： Zhu, Dan;Su, Tong;Sun, Tao;Qin, Xingcai;Su, Shao;Chao, Jie;Wang, Lianhui;Zhu, Dan;Su, Tong;Sun, Tao;Qin, Xingcai;Su, Shao;Chao, Jie;Wang, Lianhui;Li, Fang;Zhao, Dongming;Bai, Yun;Shao, Guoqing;Feng, Zhixin

作者机构：

期刊名称：ANALYTICAL CHEMISTRY （ 影响因子：7.4； 五年影响因子：7.0 ）

ISSN： 0003-2700

年卷期： 2024 年 96 卷 13 期

页码：

收录情况： SCI

摘要： Accurate, ultrasensitive, and point-of-care (POC) diagnosis of the African swine fever virus (ASFV) remains imperative to prevent its spread and limit the losses incurred. Herein, we propose a CRISPR-Cas12a-assisted triplex amplified colorimetric assay for ASFV DNA detection with ultrahigh sensitivity and specificity. The specific recognition of recombinase aided amplification (RAA)-amplified ASFV DNA could activate the Cas12a/crRNA/ASFV DNA complex, leading to the digestion of the linker DNA (bio-L1) on magnetic beads (MBs), thereby preventing its binding of gold nanoparticles (AuNPs) network. After magnetic separation, the release of AuNPs network comprising a substantial quantity of AuNPs could lead to a discernible alteration in color and significantly amplify the plasmonic signal, which could be read by spectrophotometers or smartphones. By combining the RAA, CRISPR/Cas12a-assisted cleavage, and AuNPs network-mediated colorimetric amplification together, the assay could detect as low as 0.1 copies/mu L ASFV DNA within 1 h. The assay showed an accuracy of 100% for the detection of ASFV DNA in 16 swine tissue fluid samples, demonstrating its potential for on-site diagnosis of ASFV.

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