An ultrasensitive and specific ratiometric electrochemical biosensor based on SRCA-CRISPR/Cas12a system for detection of Salmonella in food

Sensitive and specific detection of Salmonella is highly desirable to guarantee food safety and prevent food-borne disease outbreaks. Herein, we proposed a ratiometric electrochemical biosensor based on the combination of SRCA and CRISPR/Cas12a system for ultrasensitive and specific detection of Salmonella. Signal amplification is accomplished by reasonably integrating rapid SRCA amplification and the trans-cleavage capabilities of Cas12a. Accordingly, the non-specific amplification was eliminated. Compared with single signal-based electrochemical approaches, the ratiometric electrochemical measurements offer self-calibration to reduce the internal/external disturbances, thereby achieving precise analytical performance and high reliability. At this strategy, based on the ratio of the current signal of ferrocene (Fc) and methylene blue (MB) (IFc/IMB), the biosensor displayed a linear range from 5.8 fg/μL to 5.8 ng/μL and detected as low as 2.08 fg/μL of Salmonella in pure cultures. For the detection of the actual samples, this biosensor was 100% sensitivity, 97.8% specificity and 98% accuracy in comparison with the international standard. And the detection results of this biosensor were consistent with real-time fluorescent quantitative PCR (RT-qPCR), which further verified the accuracy and practical application value of the biosensor. Overall, this strategy provides a valuable platform for highly specific and ultrasensitive screening of Salmonella in food.