Photoactivatable CRISPR/Cas12a Sensors for Biomarkers Imaging and Point-of-Care Diagnostics

Abstract

In recent years, the CRISPR/Cas12a-based sensing strategy has shown significant potential for specific target detection due to its rapid and sensitive characteristics. However, the "always active" biosensors are often insufficient to manipulate nucleic acid sensing with high spatiotemporal control. It remains crucial to develop nucleic acid sensing devices that can be activated at the desired time and space by a remotely applied stimulus. Here, we integrated photoactivation with the CRISPR/Cas12a system for DNA and RNA detection, aiming to provide high spatiotemporal control for nucleic acid sensing. By rationally designing the target recognition sequence, this photoactivation CRISPR/Cas12a system could recognize HPV16 and survivin, respectively. We combined the lateral flow assay strip test with the CRISPR/Cas12a system to realize the visualization of nucleic acid cleavage signals, displaying potential instant test application capabilities. Additionally, we also successfully realized the temporary control of its fluorescent sensing activity for survivin by photoactivation in vivo, allowing rapid detection of target nucleic acids and avoiding the risk of contamination from premature leaks during storage. Our strategy suggests that the CRISPR/Cas12a platform can be triggered by photoactivation to sense various targets, expanding the technical toolbox for precise biological and medical analysis. This study represents a significant advancement in nucleic acid sensing and has potential applications in disease diagnosis and treatment.

“Three-in-One” Plasmonic Au@PtOs Nanocluster Driven Lateral Flow Assay for Multimodal Cancer Exosome Biosensing

“…Lateral flow assay (LFA) has attracted wide attention in the diagnostic field due to the advantages of rapidity, low cost, and ease-of-use. − These years, LFA has been applied in various fields including environmental pollutant analysis, , food safety surveillance, , and particularly point-of-care disease diagnosis. − However, traditional Au nanoparticles (Au NPs) based LFA can only provide limited sensitivity and quantitative ability; therefore, developing novel strategies to improve the sensitivity and quantitative capability of LFAs was urgently required. , …”

Light-activated CRISPR-Cas12a for amplified imaging of microRNA in cell cycle phases at single-cell levels

“…Nonetheless, these methods did not apply the CRISPR amplification machinery and relied on amplification pathways with limited efficiencies. Photoresponsive caged CRISPR-Cas machineries were used for in vitro amplified detection of nucleic acids ( 65 – 67 ), and their integration into cells was used for light-triggered gene-editing transformations ( 68 – 70 ) and mRNA imaging ( 71 ). Nonetheless, the photoactivated CRISPR-Cas machineries used multiple caging sites in the engineered CRISPR constituents, a feature limiting the versatility of the methods.…”