Multifunctional Photoelectrochemical Biosensor Based on ZnIn₂S₄/ZnS QDs@Au–Ag-Reversed Photocurrent of Cu-Metal–Organic Framework Coupled with CRISPR/Cas-12a-Shearing for Assay of Dual Targets

Abstract

False positives and negatives in bioanalytical assays remain a persistent problem. Herein, a multifunctional photoelectrochemical (PEC) biosensor based on ZnIn 2 S 4 (ZIS)/ZnS quantum dots (QDs)@Au−Agreversed photocurrent of Cu-metal−organic framework (MOF) coupled with CRISPR/Cas-12a-shearing was innovatively developed for assay of dual targets. First, Cu-MOF as a good PEC material shows cathodic photocurrent. Then, numerous ZIS/ZnS QDs were assembled to the Au−Ag nanoparticles (NPs) to prepare a stable and highly amplified signal probe, which can just match the energy level of Cu-MOFs and realized the polarity-reversed photocurrent of Cu-MOF for the first time. As the empty-core nanostructure of Au−Ag NPs has a high specific surface area and low material density, the bimetallic nanocrystal can much increase the reaction rate and improve the redox efficiency. When target CEA-produced cDNA opened the hairpin DNA (HP1 DNA) on the electrode, the ZIS/ZnS QDs@Au−Ag signal probe was conjugated to the electrode via DNA hybridization, achieving a significantly reversed PEC current for CEA detection. Moreover, the specific binding of kanamycin/aptamer generated the acDNA (activator), which can activate the trans-cleavage activity of the CRISPR-CAS12a system on ssDNA, so the signal probe was sheared and caused the obvious decrease of PEC signal for kanamycin detection. The newly developed ZIS/ZnS QDs@Au−Ag NPs displayed excellent PEC properties and reversed photocurrent to MOF and were combined with the unique CRISPR-Cas12a system to achieve sensitive detection of dual targets, which can open a new polarity-reversed PEC sensing platform for rapid and accurate analysis of multiple targets and can effectively avoid false positives results in clinical testing.

Low-Background CRISPR/Cas12a Sensors for Versatile Live-Cell Biosensing

“…The recent discovery and development of clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas), together known as the CRISPR/Cas system, has sparked a boom in the fields of genome editing and medical diagnosis. − Particularly, as a well-known member of the CRISPR/Cas family, CRISPR/Cas12a is a class 2 type V CRISPR-associated enzyme that binds to a single-stranded guide CRISPR RNA (crRNA). Compared with the other members of the CRISPR/Cas family, CRISPR/Cas12a has some unique characteristics. , It is able to cleave not only double-stranded DNA (dsDNA) with high sequence-specificity ( cis -cleavage activity) but also single-stranded DNA (ssDNA) in a sequence-independent manner ( trans -cleavage activity). ,− In view of the unique properties, numerous Cas12a-based biosensors have been developed for the detection of different targets. − Despite remarkable progress in recent years, − CRISPR/Cas12a-based biosensors still mainly target at nucleic acids and are rarely used in live cells. Scientists are curious about whether the CRISPR/Cas system can be deployed as a general live cell biosensing tool for a wide range of intracellular biomolecules and not just nucleic acid-associated targets.…”

Spatial-Potential-Color-Resolved Bipolar Electrode Electrochemiluminescence Biosensor Using a CuMoOx Electrocatalyst for the Simultaneous Detection and Imaging of Tetracycline and Lincomycin

“…Antibiotics are widely used in medical care, livestock breeding, agricultural production, and other industries due to their ability to inhibit and kill bacteria, making great contributions to social and economic development. − However, overuse of antibiotics can lead to high levels of toxic residues, and excessive human ingestion from the environment can cause serious side effects (such as liver and gastrointestinal system damage, anaphylactic shock, and even death). − Because the concentration of antibiotics in food and environment is relatively low, the detection of antibiotics is usually trace analysis . Therefore, it is urgent to establish a rapid, simple, low-cost, and highly specific antibiotic detection method.…”

Novel Porphyrinic Covalent Organic Polymer with Polarity-Switchable Dual Wavelength for Accurate and Sensitive Photoelectrochemical Sensing

“…The accurate and sensitive assay is crucial for life health − and thus has attracted the attention of multitudinous researchers. PEC biosensor, as an emerging and promising analysis device, has the reputation of low background and high sensitivity originating from a completely separated light excitation-electricity detection mode. − It is widely known that real samples such as serum, cells, and food contain many potential redox interferences, − so the quest for the advanced PEC biosensor with improved accuracy and sensitivity in the actual analysis remains attractive. Accordingly, the adoption of multimodal or ratiometric detection using at least two output signals has brought down.…”