Asymmetric CRISPR-Cas12a powered electrochemical aptasensor for clenbuterol detection based on competitive gRNA mediated cascade signal amplification.

The residue of clenbuterol (CLB) in food poses a potential harm to human health. Herein, we presented an electrochemical aptasensor (E-A-CRISPR) based on employing an aptamer as a specific recognition element and asymmetric CRISPR-Cas12a as signal amplifiers for sensitive, and selective detection of CLB. In this E-A-CRISPR system, the target CLB bound to the aptamer and initiated cascade signal amplification through the DNase activity of CRISPR-Cas12a with two competitive gRNAs. Upon amplification, the active Cas12a cleaved the methylene blue-labeled hairpin probe on the electrode, reducing the peak current. Under optimal conditions, the E-A-CRISPR system showed a wide linear range (1 pM-100 nM) and a low detection limit (500 fM). This system could detect CLB in potable water, pig liver, and pork samples, showing significant potential for food safety monitoring. To our knowledge, this study is the first to use a CRISPR-Cas12a powered system for electrochemical sensing of CLB.