CRISPR/Cas-powered nucleic acid amplification and amplification-free biosensors for public safety detection: Principles, advances and prospects

Rapid, accurate, cost-effective, and efficient ultrasensitive detection strategies are essential for public health safety (including food safety, disease prevention and environmental governance). The CRISPR/CRISPR-associated (Cas) detection is a cutting-edge technology that has been widely used in the detection of public health safety due to its targeted cleavage properties (signal amplification), attomolar level sensitivity, high specificity (recognizing single-base mismatches), and rapid turnover time. However, the current research about CRISPR/Cas-based biosensors is not clear, such as mechanism problem and application differences of integrating CRISPR/Cas system with other technologies, and how to further innovate and develop in the future. Therefore, further detailed analysis and comparative discussion of CRISPR/Cas-based biosensors is needed. Currently, CRISPR/Cas system powered biosensors can be mainly categorized into two types: CRISPR/Cas system powered nucleic acid amplification biosensors and CRISPR/Cas system powered nucleic acid amplification-free biosensors. The two biosensors have different characteristics and advantages. This paper first provides an in-depth investigation of the enzymatic mechanism of CRISPR/Cas system at the molecular level. Then, this paper summarizes the principles and recent advances of CRISPR/Cas system powered nucleic acid amplification biosensors and CRISPR/Cas system powered nucleic acid amplification-free biosensors and discusses their integration mechanisms in depth. More, the differences and application-oriented between the two biosensors are further discussed. Finally, the application orientation and future perspectives of the two biosensors are discussed, and unique insights into the future development of CRISPR/Cas system are provided.