Signal-amplification strategy and advances in electrochemistry-based CRISPR/Cas12 biosensing

CRISPR/Cas12 system has gained considerable attention due to its precise gene editing capabilities and robust trans-cleavage activities, demonstrating significant promise in detection of nucleic acids, proteins, ions, and microbes. The integration of CRISPR/Cas12 systems into electrochemical biosensors offers a significant opportunity to enhance adaptability, sensitivity, and specificity, reduce recovery times, has become an immensely effective tool for next-generation molecular diagnosis. To improve the performance of electrochemistry-based CRISPR/Cas12 biosensing, various signal amplification techniques have been elaborately incorporated. This review first introduces the fundamental principles underlying electrochemistry-based CRISPR/Cas12 biosensing, followed by a comprehensive analysis of advancements in two main signal amplification strategies: nucleic acid amplification-free and nucleic acid amplification-based strategies. The former one includes strategies such as magnetic nanoparticles for enhanced specificity, probe modifications to minimize background noise, crRNA design for improved efficiency, and nanomaterials-based conductivity enhancement. The latter explores innovative nucleic acid amplification techniques, including rolling circle amplification (RCA), hybridization chain reaction (HCR), and primer exchange reaction (PER) to further boost sensitivity and specificity. Finally, the challenges and future perspectives of electrochemistry-based CRISPR/Cas12 biosensing are critical examined.