Department of Central Laboratory, Shanghai Chest Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200030, P. R. China.
Research Center of Molecular Recognition and Biosensing, School of Life Sciences, Shanghai University, Shanghai 200444, P. R. China.
Anal Chem. 2022 Sep 13;94(36):12523-12530. doi: 10.1021/acs.analchem.2c02776. Epub 2022 Aug 30.
In this work, boronic ester-mediated dual recognition has been coupled with a CRISPR/Cas12a system; thus, a new method for highly specific and sensitive detection of lipopolysaccharide (LPS) is proposed via the simultaneous recognition of boronic acid and an LPS aptamer (LPSA) as well as signal amplification by CRISPR/Cas12a. Specifically, boronic acid-modified magnetic beads (MB@APBA) and aptamers are employed for the simultaneous dual recognition of LPS, while polymerase isotherm amplification is further utilized to induce LPS cycling and form a double strand, which can activate the CRISPR/Cas12a system so as to amplify the signal. Consequently, a linear detection range can be obtained from 0.05 to 5000 ng/mL, with the lowest detection limit of 44.86 pg/mL. The capturing of MB@APBA on 1, 2- and 1, 3- dihydroxyl-containing substances can not only eliminate the interference of other molecules but also enhance the highly specific recognition of LPSA on LPS. Moreover, MB@APBA can be reused by adjusting the pH value of the reaction system. The method can be developed as a universal platform for the analytical detection of other carbohydrates.
在这项工作中，硼酯介导的双重识别与 CRISPR/Cas12a 系统相结合；因此，通过同时识别硼酸和脂多糖适体（LPSA）以及通过 CRISPR/Cas12a 进行信号放大，提出了一种用于高度特异性和灵敏检测脂多糖 (LPS) 的新方法。具体来说，采用硼酸修饰的磁性珠（MB@APBA）和适体同时进行 LPS 的双重识别，同时利用聚合酶等温扩增诱导 LPS 循环并形成双链，从而激活 CRISPR/Cas12a 系统以放大信号。因此，线性检测范围可以从 0.05 到 5000 ng/mL，最低检测限为 44.86 pg/mL。含 1,2-和 1,3-二羟基的物质上的 MB@APBA 捕获不仅可以消除其他分子的干扰，还可以增强 LPS 上 LPSA 的高度特异性识别。此外，通过调整反应体系的 pH 值可以重复使用 MB@APBA。该方法可以开发为用于分析检测其他碳水化合物的通用平台。
