Abstract
COVID-19 is an ongoing pandemic caused by the highly infectious coronavirus SARS-CoV-2 that is engaging worldwide scientific research to find a timely and effective eradication strategy. Great efforts have been put into anti-COVID-19 vaccine generation in an effort to protect the world population and block SARS-CoV-2 spread. To validate the protective efficacy of the vaccination campaign and effectively control the pandemic, it is necessary to quantify the induction of neutralizing antibodies by vaccination, as they have been established to be a correlate of protection. In this work, a SARS-CoV-2 pseudovirus neutralization assay, based on a replication-incompetent lentivirus expressing an adapted form of CoV-2 S protein and an ACE2/TMPRSS2 stably expressing cell line, has been minimized in terms of protocol steps without loss of accuracy. The goal of the present simplified neutralization system is to improve SARS-CoV-2 vaccination campaign by means of an easy and accessible approach to be performed in any medical laboratory, maintaining the sensitivity and quantitative reliability of classical serum neutralization assays. Further, this assay can be easily adapted to different coronavirus variants by simply modifying the pseudotyping vector.
Highlights
Coronaviruses, belonging to the family Coronaviridae in the order Nidovirales, are positive-strand RNA viruses with a genome length of between 26 and 32 kbp
Due to its high infectivity and pathogenicity, CoV-2 needs to be handled in biosafety level 3 (BSL-3) specific facilities [10], which limits the development of antiviral measures as well as basic and applied studies on the interaction between host cells and CoV-2 and viral attachment and entry mediated by the S protein
PEF1αACE2/TMPRRS2/Puro was generated (Figure S1A). pEF1α-angiotensin-converting enzyme 2 (ACE2)/TMPRRS2/Puro was electroporated in HEK293T cells, and stably transfected cells were obtained by selection with puromycin
Summary
Gaetano Donofrio 1,*, Valentina Franceschi 1 , Francesca Macchi 1, Luca Russo 1, Anna Rocci 2, Valentina Marchica 3,4 , Federica Costa 3,4 , Nicola Giuliani 3 , Carlo Ferrari 3,5 and Gabriele Missale 3,5.
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