A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

National Science Review, Journal Year: 2022, Volume and Issue: 9(8)

Published: June 3, 2022

The outbreak of the COVID-19 pandemic was partially due to challenge identifying asymptomatic and presymptomatic carriers virus, thus highlights a strong motivation for diagnostics with high sensitivity that can be rapidly deployed. On other hand, several concerning SARS-CoV-2 variants, including Omicron, are required identified as soon samples 'positive'. Unfortunately, traditional PCR test does not allow their specific identification. Herein, first time, we have developed MOPCS (Methodologies Photonic CRISPR Sensing), which combines an optical sensing technology-surface plasmon resonance (SPR) 'gene scissors' clustered regularly interspaced short palindromic repeat (CRISPR) technique achieve both specificity when it comes measurement viral variants. is low-cost, CRISPR/Cas12a-system-empowered SPR gene-detecting platform analyze RNA, without need amplification, within 38 min from sample input results output, limit detection 15 fM. achieves highly sensitive analysis SARS-CoV-2, mutations appear in variants B.1.617.2 (Delta), B.1.1.529 (Omicron) BA.1 (a subtype Omicron). This also used some recently collected patient local China, by Centers Disease Control Prevention. innovative CRISPR-empowered will further contribute fast, accurate target nucleic acid sequences single-base mutations.

Language: Английский

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A Highly Sensitive CRISPR‐Empowered Surface Plasmon Resonance Sensor for Diagnosis of Inherited Diseases with Femtomolar‐Level Real‐Time Quantification

Advanced Science, Journal Year: 2022, Volume and Issue: 9(14)

Published: March 27, 2022

The clustered regularly interspaced short palindromic repeats (CRISPR) molecular system has emerged as a promising technology for the detection of nucleic acids. Herein, development surface plasmon resonance (SPR) sensor that is functionalized with layer locally grown graphdiyne film, achieving excellent sensing performance when coupled catalytically deactivated CRISPR-associated protein 9 (dCas9), reported. dCas9 immobilized on and complexed specific single-guide RNA, enabling amplification-free target sequences within genomic DNA. sensor, termed CRISPR-SPR-Chip, used to successfully analyze recombinant plasmids only three-base mutations limit low 1.3 fM. Real-time monitoring CRISPR-SPR-Chip clinical samples patients Duchenne muscular dystrophy two exon deletions, which are detected without any pre-amplification step, yielding significantly positive results 5 min. ability this novel CRISPR-empowered SPR (CRISPR-eSPR) platform rapidly, precisely, sensitively, specifically detect gene sequence provides new on-chip optic approach analysis.

Language: Английский

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The CRISPR–Cas toolbox for analytical and diagnostic assay development

Chemical Society Reviews, Journal Year: 2021, Volume and Issue: 50(21), P. 11844 - 11869

Published: Jan. 1, 2021

A comprehensive review that offers mechanistic insight into the CRISPR–Cas toolbox for analytical and diagnostic assay development.

Language: Английский

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Size-Controlled Engineering Photoelectrochemical Biosensor for Human Papillomavirus-16 Based on CRISPR-Cas12a-Induced Disassembly of Z-Scheme Heterojunctions

ACS Sensors, Journal Year: 2022, Volume and Issue: 7(5), P. 1593 - 1601

Published: May 5, 2022

Photoelectrochemical (PEC) biosensors incorporating biomolecular recognition with photon-to-electron conversion capabilities of the photoactive species have been developed for molecular diagnosis, but most involve difficulty in adjusting band gap positions and are unsuitable PEC biodetection. In this work, an innovative biosensor combined quantum size-controlled engineering based on confinement by controlling size was designed detection human papillomavirus-16 (HPV-16) through CRISPR-Cas12a (Cpf1)-induced disassembly Z-scheme heterojunction. To best our knowledge, that precisely tunes properties materials is first utilized bioanalysis. Based effect, light absorption efficiency charge-transfer rate were tuned to suitable levels obtain performance. After incubation target HPV-16, binding Cas12a-crRNA double-stranded DNA (dsDNA) stimulated activity indiscriminate cleavage toward single-stranded (ssDNA), resulting a decrease photocurrent due blocking electron transfer By optimizing experimental conditions, sensing system exhibited incredible response HPV-16 range from 3.0 pM 600 nM limit 1.0 pM. Impressively, application effect could stimulate more interest precise design structure improve

Language: Английский

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Universal and Programmable Rolling Circle Amplification-CRISPR/Cas12a-Mediated Immobilization-Free Electrochemical Biosensor

Analytical Chemistry, Journal Year: 2021, Volume and Issue: 93(20), P. 7499 - 7507

Published: May 13, 2021

The development of a sensing platform with high sensitivity and specificity, especially programmability universal applicability, for the detection clinically relevant molecules is highly valuable disease monitoring confirmation but remains challenge. Here, first time, we introduce clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system into an immobilization-free electrochemical biosensing sensitively specifically detecting disease-related nucleic acids small molecules. In this strategy, modular rolling circle amplification (RCA) designed to transform amplify target recognition event trigger DNA strand that used as activate deoxyribonuclease activity CRISPR/Cas12a further signal amplification. cleavage target-activated blocker probe allows methylene blue-labeled reporter probes be captured by reduced graphene oxide-modified electrode, leading obviously increased signal. We only need simply tune sequence in RCA components, strategy can flexibly applied sensitive specific microRNAs, Parvovirus B19 DNA, adenosine-5′-triphosphate calculated limit 0.83 aM, 0.52 0.46 pM, respectively. addition, construct logic circuits (YES, NOT, OR, AND) inputs experimentally demonstrate modularity stimuli-responsive RCA-CRISPR/Cas12a system. This work broadens application provides new thinking developing robust tool clinical diagnosis.

Language: Английский

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Photoelectrochemical bioanalysis of microRNA on yolk-in-shell Au@CdS based on the catalytic hairpin assembly-mediated CRISPR-Cas12a system

Chemical Communications, Journal Year: 2022, Volume and Issue: 58(54), P. 7562 - 7565

Published: Jan. 1, 2022

This work reports on the proof-of-concept of a photoelectrochemical (PEC) biosensor with horseradish peroxidase-single stranded DNA-encoded magnetic bead (MB-ssDNA-HRP) signal probe cleaved by catalytic hairpin assembly (CHA)-mediated clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a system for quantification microRNA (miR-21) using yolk-in-shell Au@CdS as photoactive material.

Language: Английский

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Cas12a-based electrochemiluminescence biosensor for target amplification-free DNA detection

Biosensors and Bioelectronics, Journal Year: 2021, Volume and Issue: 176, P. 112954 - 112954

Published: Jan. 1, 2021

Language: Английский

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Exploring the Trans-Cleavage Activity of CRISPR/Cas12a on Gold Nanoparticles for Stable and Sensitive Biosensing

Analytical Chemistry, Journal Year: 2021, Volume and Issue: 93(11), P. 4967 - 4974

Published: March 11, 2021

Taking advantage of the excellent trans-cleavage activity, CRISPR-based diagnostics (CRISPR-Dx) has shown great promise in molecular diagnostics. However, single-stranded DNA reporter current CRISPR-Dx suffers from poor stability and limited sensitivity, which make their application complex biological environments difficult. Herein, we, for first time, explore activity CRISPR/Cas12a toward substrate on gold nanoparticles apply new phenomenon to develop a spherical nucleic acid (SNA) stable sensitive biosensing. By anchoring nanoparticles, we discovered different activities types Cas12a system (e.g., LbCas12a AsCas12a) nanoparticle surface. The further study suggests that surface is highly dependent density length strands. Based these interesting discoveries, furthermore SNA reporter-based fluorescent biosensing application. Compared traditional ssDNA reporters, exhibits improved stability, enables serum environment. In addition, with tunable high sensitivity detection limit 10 fM, about 2 orders magnitude lower than system. Finally, practical clinical was successfully achieved. These results indicate significant potential future research biology science medical diagnoses.

Language: Английский

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CRISPR Enzyme Kinetics for Molecular Diagnostics

Analytical Chemistry, Journal Year: 2021, Volume and Issue: 93(20), P. 7456 - 7464

Published: May 12, 2021

CRISPR-diagnostic assays have gained significant interest in the last few years. This has grown rapidly during current COVID-19 pandemic, where CRISPR-diagnostics been frontline contenders for rapid testing solutions. surge research prompts following question: what exactly are achievable limits of detection and associated assay times enabled by kinetics enzymes such as Cas12 Cas13? To explore this question, we here present a model based on Michaelis–Menten enzyme theory applied to CRISPR enzymes. We use develop analytical solutions reaction back-of-the-envelope criteria validate check consistency reported kinetic parameters. our analyses all studies known us, which report Michaelis–Menten-type data CRISPR-associated These include subtypes Cas13 orthologs. found but one study clearly violate at least two three rules therefore that basic physical limits. performed an experimental LbCas12a with both ssDNA dsDNA activators these its predicted scaling. The validated is used time scales degree completion practically relevant target concentrations applicable assays. results broad implications emerging, amplification-free CRISPR-detection methods.

Language: Английский

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CRISPR/Cas12a-based photoelectrochemical sensing of microRNA on reduced graphene oxide-anchored Bi2WO6 coupling with catalytic hairpin assembly

Sensors and Actuators B Chemical, Journal Year: 2022, Volume and Issue: 369, P. 132307 - 132307

Published: July 5, 2022

Language: Английский

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