Reagents-Loaded, Automated Assay that Integrates Recombinase-Aided Amplification and Cas12a Nucleic Acid Detection for a Point-of-Care Test
Yong Chen, Yixin Mei, Xiaohui Zhao, Xingyu Jiang
Abstract
Point-of-care testing for nucleic acid that combines amplification and readout is in great demand. This report integrates recombinase-aided amplification (RAA) with the clustered regularly interspaced short palindromic repeats (CRISPR)–Cas12a system in a centrifugal microfluidic point-of-care test. We overcome the difficulty in the integration of these two processes, which mainly lies in the fact that Cas12a can digest the template DNA. We further integrate all reagents into the centrifugal microfluidics to automate the entire process. The Cas12a-assisted straightforward microfluidic equipment for analysis of nucleic acid (CASMEAN) enables us to rapidly and conveniently detect nucleic acid within 1.5 h. An application for detecting Pseudomonas aeruginosa confirms the excellent compatibility between RAA and the Cas12a system, which ensures the superior performance of CASMEAN, such as capabilities of point-of-care detection, high sensitivity, and high specificity. CASMEAN is a genetic detection platform with great potential.