CRISPR-Cas12a-Based Aptasensor for On-Site and Highly Sensitive Detection of Microcystin-LR in Freshwater

On-site monitoring of trace organic pollutants with facile methods is critical to environmental pollutant prevention and control. Herein, we proposed a CRISPR-Cas12a-based aptasensor platform (named as MC-LR-Casor) for on-site and sensitive detection of microcystin-LR (MC-LR). After hybridization with blocker DNA, the MC-LR aptamers were conjugated to magnetic beads (MBs) to get the MB aptasensor. In the presence of MC-LR, their interactions with aptamers were triggered and the specific binding caused the release of blocker DNA. Using the programmability of the CRISPR-Cas system, the released blocker DNA was designed to activate a Cas12a-crRNA complex. Single strand DNA reporters were rapidly cleaved by the complex Signal readout could be achieved by fluorometer or lateral flow strips, which were positively correlated to MC-LR concentration. Benefiting from the CRISPR-Cas12a amplification system, the proposed sensing platform exhibited high sensitivity and reached the limit of detection of similar to 3 x 10(-6) mu g/L (fluorescence method) or 1 x 10(-3) mu g/L (lateral flow assay). In addition, the MC-LR-Casor showed excellent selectivity and good recovery rates, demonstrating their good applicability for real water sample analysis. During the whole assay, only two steps of incubation at a constant temperature were required and the results could be visualized when employing flow strips. Therefore, the proposed assay offered a simple and convenient alternative for in situ MC-LR monitoring, which may hold great promise for future environmental surveillance.

Automated summary

In this study, a CRISPR-Cas12a-based aptasensor platform was proposed for on-site detection of microcystin-LR (MC-LR). The platform combined magnetic beads and aptamers to achieve signal output, utilizing the programmability of the CRISPR-Cas system. Successful detection of MC-LR in water samples was demonstrated.