Paper-Supported Photoelectrochemical Biosensor for Dual-Mode miRNA-106a Assay: Integration of Luminescence-Confined Upconversion-Actuated Fluorescent Resonance Energy Transfer and CRISPR/Cas13a-Powered Cascade DNA Circuits

Near-infrared (NIR)-responsive bioassays based on upconversion nanoparticle (UCNP) incorporating high-performance semiconductors have been developed by researchers, but most lack satisfactory ultrasensitivity for exceedingly trace amounts of target. Herein, for the first time, the CRISPR/Cas13a system is combined with cascade DNA circuits, fluorescent resonance energy transfer (FRET) effect, and luminescence-confined UCNPs-bonded CuInS2/ZnO p-n heterostructures-functionalized paper-working electrode to construct dual-signal-on paper-supported NIR-irradiated photoelectrochemical (PEC) (NIR-PEC) and upconversion luminescence (UCL) bioassay for high-sensitive quantification of miRNA-106a (miR-106a). By constructing an ideal FAM-labeled aminating molecular beacon (FAM-H2) model, a relatively good FRET ratio between the UCNP and FAM (approximate to 85.3%) can be achieved. In the existence of miR-106a, the hairpin-structure FAM-H2 was unwound, bringing about the distance increase of UCNP and FAM and the restraint of FRET. Accordingly, both the NIR-PEC signal and the UCL intensity gradually recovered distinctly. Unlike conventional single-mode PEC sensors, with NIR excitation, the designed dual-mode sensing system could implement minimized misdiagnose assay and quantitative miR-106a determination with low detection limits, that is, 76.54 and 51.36 aM for NIR-PEC and UCL detection, respectively. This work not only broadens the horizon of application of the CRISPR/Cas13a strategy toward biosensing but also constructs a new structure of the UCNP-semiconductor in the exploration of efficient NIR-responsive tools and inspires the construction of a no-misdiagnosed and novel biosensor for dual-mode liquid biopsy.

Automated summary

Researchers have developed a near-infrared-responsive bioassay for high-sensitive quantification of miRNA-106a by combining the CRISPR/Cas13a system with cascade DNA circuits, FRET effect, and UCNP-semiconductor. This dual-signal-on sensing system enables minimized misdiagnose assay and has low detection limits.