Rapid Isothermal Amplification and Portable Detection System for SARS-CoV-2
The COVID-19 pandemic provides an urgent example where a gap exists between availability of state-of-the-art diagnostics and current needs. As assay details and primer sequences become widely known, many laboratories could perform diagnostic tests using methods such as RT-PCR or isothermal RT-LAMP amplification. A key advantage of RT-LAMP based approaches compared to RT-PCR is that RT-LAMP is known to be robust in detecting targets from unprocessed samples. In addition, RT-LAMP assays are performed at a constant temperature enabling speed, simplicity, and point-of-use testing. Here, we provide the details of an RT-LAMP isothermal assay for the detection of SARS-CoV-2 virus with performance comparable to currently approved tests using RT-PCR. We characterize the assay by introducing swabs in virus spiked synthetic nasal fluids, moving the swab to viral transport medium (VTM), and using a volume of that VTM for performing the amplification without an RNA extraction kit. The assay has a Limit-of-Detection (LOD) of 50 RNA copies/μL in the VTM solution within 20 minutes, and LOD of 5000 RNA copies/μL in the nasal solution. Additionally, we show the utility of this assay for real-time point-of-use testing by demonstrating detection of SARS-CoV-2 virus in less than 40 minutes using an additively manufactured cartridge and a smartphone-based reader. Finally, we explore the speed and cost advantages by comparing the required resources and workflows with RT-PCR. This work could accelerate the development and availability of SARS-CoV-2 diagnostics by proving alternatives to conventional laboratory benchtop tests.
| Errataetall: |
UpdateIn: Proc Natl Acad Sci U S A. 2020 Aug 31;:. - PMID 32868442 |
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| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2020 |
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| Erschienen: |
2020 |
| Enthalten in: |
Zur Gesamtaufnahme - year:2020 |
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| Enthalten in: |
bioRxiv : the preprint server for biology - (2020) vom: 21. Mai |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Ganguli, A [VerfasserIn] |
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| Links: |
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| Themen: |
Additive manufacturing |
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| Anmerkungen: |
Date Revised 16.02.2024 published: Electronic UpdateIn: Proc Natl Acad Sci U S A. 2020 Aug 31;:. - PMID 32868442 Citation Status PubMed-not-MEDLINE |
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| doi: |
10.1101/2020.05.21.108381 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM310908280 |
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| 520 | |a The COVID-19 pandemic provides an urgent example where a gap exists between availability of state-of-the-art diagnostics and current needs. As assay details and primer sequences become widely known, many laboratories could perform diagnostic tests using methods such as RT-PCR or isothermal RT-LAMP amplification. A key advantage of RT-LAMP based approaches compared to RT-PCR is that RT-LAMP is known to be robust in detecting targets from unprocessed samples. In addition, RT-LAMP assays are performed at a constant temperature enabling speed, simplicity, and point-of-use testing. Here, we provide the details of an RT-LAMP isothermal assay for the detection of SARS-CoV-2 virus with performance comparable to currently approved tests using RT-PCR. We characterize the assay by introducing swabs in virus spiked synthetic nasal fluids, moving the swab to viral transport medium (VTM), and using a volume of that VTM for performing the amplification without an RNA extraction kit. The assay has a Limit-of-Detection (LOD) of 50 RNA copies/μL in the VTM solution within 20 minutes, and LOD of 5000 RNA copies/μL in the nasal solution. Additionally, we show the utility of this assay for real-time point-of-use testing by demonstrating detection of SARS-CoV-2 virus in less than 40 minutes using an additively manufactured cartridge and a smartphone-based reader. Finally, we explore the speed and cost advantages by comparing the required resources and workflows with RT-PCR. This work could accelerate the development and availability of SARS-CoV-2 diagnostics by proving alternatives to conventional laboratory benchtop tests | ||
| 650 | 4 | |a Preprint | |
| 650 | 4 | |a Additive manufacturing | |
| 650 | 4 | |a COVID19 | |
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| 650 | 4 | |a SARS-CoV-2 | |
| 650 | 4 | |a Viral detection | |
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| 700 | 1 | |a Sun, F |e verfasserin |4 aut | |
| 700 | 1 | |a Valera, E |e verfasserin |4 aut | |
| 700 | 1 | |a Cunningham, B T |e verfasserin |4 aut | |
| 700 | 1 | |a King, W P |e verfasserin |4 aut | |
| 700 | 1 | |a Bashir, R |e verfasserin |4 aut | |
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