Ultra-sensitive and high-throughput CRISPR-p owered COVID-19 diagnosis

Recent research suggests that SARS-CoV-2-infected individuals can be highly infectious while asymptomatic or pre-symptomatic, and that an infected person may infect 5.6 other individuals on average. This situation highlights the need for rapid, sensitive SARS-CoV-2 diagnostic assays capable of high-...

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Veröffentlicht in:Biosensors & bioelectronics Jg. 164; S. 112316
Hauptverfasser: Huang, Zhen, Tian, Di, Liu, Yang, Lin, Zhen, Lyon, Christopher J., Lai, Weihua, Fusco, Dahlene, Drouin, Arnaud, Yin, Xiaoming, Hu, Tony, Ning, Bo
Format: Journal Article
Sprache:Englisch
Veröffentlicht: England Elsevier B.V 15.09.2020
Schlagworte:
Base Sequence
Betacoronavirus - genetics
Betacoronavirus - isolation & purification
Biosensing Techniques - methods
Biosensing Techniques - statistics & numerical data
biosensors
Coronavirus infections
Coronavirus Infections - diagnosis
Coronavirus Infections - virology
COVID-19
CRISPR
CRISPR-Cas Systems
detection limit
diagnostic techniques
fluorescence
Fluorescent detection
Fluorescent Dyes
gene editing
Genes, Viral
High-Throughput Nucleotide Sequencing - methods
High-Throughput Nucleotide Sequencing - statistics & numerical data
Highly sensitive
Humans
Molecular diagnosis
nose
Nucleic Acid Amplification Techniques - methods
Nucleic Acid Amplification Techniques - statistics & numerical data
Pandemics
Pneumonia, Viral - diagnosis
Pneumonia, Viral - virology
Predictive Value of Tests
quantitative polymerase chain reaction
recombinases
Reproducibility of Results
reverse transcriptase polymerase chain reaction
Reverse Transcriptase Polymerase Chain Reaction - methods
Reverse Transcriptase Polymerase Chain Reaction - statistics & numerical data
RNA, Viral - analysis
RNA, Viral - genetics
SARS-CoV-2
screening
Sensitivity and Specificity
COVID-19
Fluorescent detection
CRISPR
SARS-CoV-2
Molecular diagnosis
Highly sensitive
ISSN:0956-5663, 1873-4235, 1873-4235
Online-Zugang:Volltext
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Zusammenfassung:Recent research suggests that SARS-CoV-2-infected individuals can be highly infectious while asymptomatic or pre-symptomatic, and that an infected person may infect 5.6 other individuals on average. This situation highlights the need for rapid, sensitive SARS-CoV-2 diagnostic assays capable of high-throughput operation that can preferably utilize existing equipment to facilitate broad, large-scale screening efforts. We have developed a CRISPR-based assay that can meet all these criteria. This assay utilizes a custom CRISPR Cas12a/gRNA complex and a fluorescent probe to detect target amplicons produced by standard RT-PCR or isothermal recombinase polymerase amplification (RPA), to allow sensitive detection at sites not equipped with real-time PCR systems required for qPCR diagnostics. We found this approach allowed sensitive and robust detection of SARS-CoV-2 positive samples, with a sample-to-answer time of ~50 min, and a limit of detection of 2 copies per sample. CRISPR assay diagnostic results obtained nasal swab samples of individuals with suspected COVID-19 cases were comparable to paired results from a CDC-approved quantitative RT-PCR (RT-qPCR) assay performed in a state testing lab, and superior to those produced by same assay in a clinical lab, where the RT-qPCR assay exhibited multiple invalid or inconclusive results. Our assay also demonstrated greater analytical sensitivity and more robust diagnostic performance than other recently reported CRISPR-based assays. Based on these findings, we believe that a CRISPR-based fluorescent application has potential to improve current COVID-19 screening efforts. •Integrated CRISPR for COVID-19 clinical diagnosis.•Rapidly identify SARS-CoV-2-specific RNA signatures at an ultra-low concentration.•Detection environment friendly and compatible for high throughput COVID-19 screening in hospital.•Successful development and validation with swab samples from patients.•Improve diagnosis of individuals with suspected COVID-19 infections.
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ISSN:0956-5663
1873-4235
1873-4235
DOI:10.1016/j.bios.2020.112316