An ultrafast SARS-CoV-2 virus enrichment and extraction method compatible with multiple modalities for RNA detection

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a zoonotic RNA virus characterized by high transmission rates and pathogenicity worldwide. Continued control of the COVID-19 pandemic requires the diversification of rapid, easy to use, sensitive, and portable methods for SARS-CoV-2 sam...

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Bibliographic Details
Published in:Analytica chimica acta 2021-10, Vol.1180, p.338846-338846, Article 338846
Main Authors: Dignan, Leah M., Turiello, Rachelle, Layne, Tiffany R., O'Connell, Killian C., Hickey, Jeff, Chapman, Jeff, Poulter, Melinda D., Landers, James P.
Format: Article
Language:English
Subjects:
Affinity nanoparticles
COVID-19
COVID-19 Testing
Enzymatic extraction
Humans
Loop-mediated amplification (LAMP)
Molecular Diagnostic Techniques
Nucleic Acid Amplification Techniques
Pandemics
Polymerase chain reaction (PCR)
Recombinase polymerase amplification (RPA)
RNA, Viral - genetics
SARS-CoV-2
Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)
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Summary:Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a zoonotic RNA virus characterized by high transmission rates and pathogenicity worldwide. Continued control of the COVID-19 pandemic requires the diversification of rapid, easy to use, sensitive, and portable methods for SARS-CoV-2 sample preparation and analysis. Here, we propose a method for SARS-CoV-2 viral enrichment and enzymatic extraction of RNA from clinically relevant matrices in under 10 min. This technique utilizes affinity-capture hydrogel particles to concentrate SARS-CoV-2 from solution, and leverages existing PDQeX technology for RNA isolation. Characterization of our method is accomplished with reverse transcription real-time polymerase chain reaction (RT-PCR) for relative, comparative RNA detection. In a double-blind study analyzing viral transport media (VTM) obtained from clinical nasopharyngeal swabs, our sample preparation method demonstrated both comparable results to a routinely used commercial extraction kit and 100% concordance with laboratory diagnoses. Compatibility of eluates with alternative forms of analysis was confirmed using microfluidic RT-PCR (μRT-PCR), recombinase polymerase amplification (RPA), and loop-mediated isothermal amplification (LAMP). The alternative methods explored here conveyed successful amplification from all RNA eluates originating from positive clinical samples. Finally, this method demonstrated high performance within a saliva matrix across a broad range of viral titers and dilutions up to 90% saliva matrix, and sets the stage for miniaturization to the microscale. [Display omitted] •SARS-CoV-2 characterized by high transmission rates and pathogenicity worldwide.•Diversification of sample preparation methods for SARS-CoV-2 is of paramount importance.•Method for viral enrichment and enzymatic extraction of RNA in under 10 min.•Eluates are compatible with downstream real-time PCR, LAMP, and RPA.
ISSN:0003-2670
1873-4324
1873-4324
DOI:10.1016/j.aca.2021.338846