Engineered CRISPR/Cas Ribonucleoproteins for Enhanced Biosensing and Bioimaging

CRISPR-Cas systems represent a highly programmable and precise nucleic acid-targeting platform, which has been strategically engineered as a versatile toolkit for biosensing and bioimaging applications. Nevertheless, their analytical performance is constrained by inherent functional and activity limitations of natural CRISPR/Cas systems, underscoring the critical role of molecular engineering in enhancing their capabilities. This review comprehensively examines recent advancements in engineering CRISPR/Cas ribonucleoproteins (RNPs) to enhance their functional capabilities for advanced molecular detection and cellular imaging. We explore innovative strategies for developing enhanced CRISPR/Cas RNPs, including Cas protein engineering through protein mutagenesis and fusion techniques, and guide RNA engineering via chemical and structural modifications. Furthermore, we evaluate these engineered RNPs' applications in sensitive biomarker detection and live-cell genomic DNA and RNA monitoring, while analyzing the current challenges and prospective developments in CRISPR-Cas RNP engineering for advanced biosensing and bioimaging.