Chemiluminescence Sensor for miRNA-21 Detection Based on CRISPR-Cas12a and Cation Exchange Reaction

Abstract

Herein, a chemiluminescence (CL) biosensor based on CRISPR-Cas12a and cation exchange reaction was constructed to detect the biomarker microRNA-21 (miRNA-21). The rolling circle amplification (RCA) reaction was introduced to convert each target RNA strand into a long single-stranded DNA with repeated sequences, which acted as triggers to initiate the transcleavage activity of CRISPR-Cas12a. The activated Cas12a could cleave the biotinylated linker DNA of CuS nanoparticles (NPs) to inhibit the binding of CuS NPs to streptavidin immobilized on the surface of the microplate, which strongly reduced the generation of Cu2+ from a cation exchange between CuS NPs and AgNO3, and thus efficiently suppressed the CL of Cu2+-luminol-H2O2 system, giving a “signal off” biosensor. With the multiple amplification, the detection limit of the developed sensor for miRNA-21 reached 16 aM. In addition, this biosensor is not only suitable for a professional chemiluminescence instrument but also for a smartphone used as a detection tool for the purpose of portable and low-cost assay. This method could be used to specifically detect quite a low level of miRNA-21 in human serum samples and various cancer cells, indicating its potential in ultrasensitive molecular diagnostics.

Pyroelectric-Effect-Assisted Near-Infrared-Driven Photoelectrochemical Biosensor Based on Exponential DNA Amplifier for MicroRNA Detection

“…In addition, the total RNA extracted from two human cancer cell lines, cervical cancer cells (HeLa) and breast cancer cells (MCF-7) (1 × 10 6 cells for each), were analyzed (Figure H). A higher level of miRNA-21 in total extracts is presented in MCF-7 cells than in HeLa cells, which is consistent with previous literature. , Besides, the miRNA-21 content in extracts detected by the proposed biosensor is in accordance with the results obtained using a qRT-PCR kit (Figures I and S8). All the results prove the good practicability of the proposed pyroelectric-effect-assisted NIR PEC biosensor for miRNA-21 detection in complex biological systems, which is practical to the sensitive and specific analysis of miRNA in clinical applications.…”

Cation Exchange Reaction-Mediated Photothermal and Polarity-Switchable Photoelectrochemical Dual-Readout Biosensor

“…Cation exchange (CE) reaction, a process of replacing host cations of parent ionic material with foreign mobile cations, has been developed as a newly emerged technique of nanocrystal controllable synthesis for solving the challenges existing in traditional one-step synthesis to obtain nanocrystals with controllable constituents and required structures. , So far, metal-sulfide-based CE strategies have been applied in many fields containing sensing fields. For example, Zhou et al established a “signal-off” biosensor for miRNA-21 detection through decreasing Cu 2+ created by CE between CuS nanoparticles and AgNO 3 , inhibiting the CL of the Cu 2+ -luminol-H 2 O 2 system . Zhou et al designed a PEC platform based on Zn-to-Hg ion exchange for the in situ formation of double type-II heterostructures (HgS/ZnS/CdS) for Hg 2+ monitoring .…”

Ultrasensitive Detection of miRNA via CRISPR/Cas12a Coupled with Strand Displacement Amplification Reaction

“…Owing to its characteristic of highly efficient collateral cleavage, the CRISPR/Cas12a system is often used as a tool for signal amplification in biosensors. It has been extensively used for detection of nucleic acids, − metal ions, , and proteins , and shows great promise in the development of POC diagnostic devices. In addition, many amplification techniques such as polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP), catalytic hairpin assembly (CHA), hybridization chain reaction (HCR), and recombinase polymerase amplification (RPA) have been reported to be combined with the CRISPR/Cas12a system to further improve the sensitivity.…”