State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing 100071, China.
Chinese PLA Center for Disease Control and Prevention, Beijing 102206, China.
Biosensors (Basel). 2023 May 31;13(6):597. doi: 10.3390/bios13060597.
The rapid and ongoing spread of the coronavirus disease (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emphasizes the urgent need for an easy and sensitive virus detection method. Here, we describe an immunocapture magnetic bead-enhanced electrochemical biosensor for ultrasensitive SARS-CoV-2 detection based on clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins, collectively known as CRISPR-Cas13a technology. At the core of the detection process, low-cast and immobilization-free commercial screen-printed carbon electrodes are used to measure the electrochemical signal, while streptavidin-coated immunocapture magnetic beads are used to reduce the background noise signal and enhance detection ability by separating the excessive report RNA, and a combination of isothermal amplification methods in the CRISPR-Cas13a system is used for nucleic acid detection. The results showed that the sensitivity of the biosensor increased by two orders of magnitude when the magnetic beads were used. The proposed biosensor required approximately 1 h of overall processing time and demonstrated an ultrasensitive ability to detect SARS-CoV-2, which could be as low as 1.66 aM. Furthermore, owing to the programmability of the CRISPR-Cas13a system, the biosensor can be flexibly applied to other viruses, providing a new approach for powerful clinical diagnostics.
新型冠状病毒(COVID-19)是由严重急性呼吸系统综合征冠状病毒 2(SARS-CoV-2)引起的,其传播迅速且仍在持续,这凸显了对简便、灵敏的病毒检测方法的迫切需求。在这里,我们描述了一种基于成簇规律间隔短回文重复序列(CRISPR)和 CRISPR 相关(Cas)蛋白(统称为 CRISPR-Cas13a 技术)的免疫捕获磁珠增强电化学生物传感器,用于超灵敏 SARS-CoV-2 检测。在检测过程的核心部分,使用低成本、无固定化的商用丝网印刷碳电极来测量电化学信号,而链霉亲和素包被的免疫捕获磁珠用于通过分离过多的报告 RNA 来降低背景噪声信号并增强检测能力,并在 CRISPR-Cas13a 系统中结合等温扩增方法进行核酸检测。结果表明,使用磁珠后,生物传感器的灵敏度提高了两个数量级。该生物传感器总共需要大约 1 小时的处理时间,并且表现出超灵敏的 SARS-CoV-2 检测能力,其检测下限低至 1.66 aM。此外,由于 CRISPR-Cas13a 系统的可编程性,该生物传感器可以灵活应用于其他病毒,为强大的临床诊断提供了一种新方法。