Direct Lysis RT-qPCR of SARS-CoV-2 in Cell Culture Supernatant Allows for Fast and Accurate Quantification

Studying the entire virus replication cycle of SARS-CoV-2 is essential to identify the host factors involved and treatments to combat infection. Quantification of released virions often requires lengthy procedures, whereas quantification of viral RNA in supernatant is faster and applicable to clinic...

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Bibliographic Details
Published in:Viruses 2022-02, Vol.14 (3), p.508
Main Authors: Craig, Nicky, Fletcher, Sarah L, Daniels, Alison, Newman, Caitlin, O'Shea, Marie, Tan, Wenfang Spring, Warr, Amanda, Tait-Burkard, Christine
Format: Article
Language:English
Subjects:
Cell culture
Cell Culture Techniques
Clinical isolates
coronavirus
Coronaviruses
COVID-19
COVID-19 - diagnosis
Dengue fever
diagnostics
direct lysis
Drugs
Efficiency
Epidemics
Heat
High-throughput screening
Humans
Infections
Influenza
Lysis
Methods
Optimization
Pandemics
Patients
real-time PCR
Real-Time Polymerase Chain Reaction
Ribonucleic acid
RNA
RNA, Viral - analysis
SARS-CoV-2
SARS-CoV-2 - genetics
Severe acute respiratory syndrome coronavirus 2
Virions
Viruses
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Summary:Studying the entire virus replication cycle of SARS-CoV-2 is essential to identify the host factors involved and treatments to combat infection. Quantification of released virions often requires lengthy procedures, whereas quantification of viral RNA in supernatant is faster and applicable to clinical isolates. Viral RNA purification is expensive in terms of time and resources, and is often unsuitable for high-throughput screening. Direct lysis protocols were explored for patient swab samples, but the lack of virus inactivation, cost, sensitivity, and accuracy is hampering their application and usefulness for in vitro studies. Here, we show a highly sensitive, accurate, fast, and cheap direct lysis RT-qPCR method for quantification of SARS-CoV-2 in culture supernatant. This method inactivates the virus and permits detection limits of 0.043 TCID50 virus and
ISSN:1999-4915
1999-4915
DOI:10.3390/v14030508