Holographic Optical Tweezers and Boosting Upconversion Luminescent Resonance Energy Transfer Combined Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas12a Biosensors

Taking advantage of outstanding precision in target recognition and trams-cleavage ability, the recently discovered CRISPR/Cas12a system provides an alternative opportunity for designing fluorescence biosensors. To fully exploit the analytical potential, we introduce here some meaningful concepts. First, the collateral cleavage of CRISPR/Cas12a is efficiently activated in a functional DNA regulation manner and the bottleneck which largely applicable to nucleic acids detection is broken. After selection of a representative aptamer and DNAzyme as the transduction pathways, the sensing coverage is extended to a small organic compound (ATP) and a metal ion (Na+). The assay sensitivity is significantly improved by utilizing a bead-supported enrichment strategy wherein emerging holographic optical tweezers are used to enhance imaging stability and simultaneously achieve multiflux analysis. Last, a sandwich-structured energy-concentrating upconversion nanoparticle triggered boosting luminescent resonance energy transfer mode is comined to face with complicated biological samples by skillfully confining the emitters into a very limited inner shell. Following the above attempts, the developed CRISPR/Cas12a biosensors not only present an ultrasensitive assay behavior toward these model non-nucleic acid analytes but also can serve as a formidable toolbox for determining real samples including single cell lysates and human plasma, proving a good practical application capacity.

Automated summary

The CRISPR/Cas12a system offers an alternative opportunity for designing fluorescence biosensors with outstanding precision in target recognition. By utilizing collateral cleavage and representative transduction pathways, the system can extend sensing coverage and improve assay sensitivity. The developed biosensors not only exhibit ultrasensitive behavior towards non-nucleic acid analytes but also show practical application capacity in real samples.