Production, titration, neutralisation, storage and lyophilisation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lentiviral pseudotypes

Authors: Di Genova, Cecilia;

Sampson, Alex;

Scott, Simon;

Cantoni, Diego;

Neto, Martin;

Bentley, Emma M.;

Mattiuzzo, Giada;

Wright, Edward;

Derveni, Mariliza;

Auld, Bethany;

Ferrara, Bill;

Harrison, Dale;

Said, Mohamed;

Selim, Arwa;

Thompson, Elinor;

Thompson, Craig P. ;

Carnell, George W.;

Temperton, Nigel

Source: Bio-Protocol Journal

Full text

: https://doi.org/10.21769/BioProtoc.4236

Abstract

This protocol details a rapid and reliable method for the production and titration of high-titre viral pseudotype particles with the SARS-CoV-2 spike protein (and D614G or other variants of concern, VOC) on a lentiviral vector core, and use for neutralisation assays in target cells expressing angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). It additionally provides detailed instructions on substituting in new spike variants via gene cloning, lyophilisation and storage/shipping considerations for wide deployment potential. Results obtained with this protocol show that SARS-CoV-2 pseudotypes can be produced at equivalent titres to SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV) pseudotypes, neutralised by human convalescent plasma and monoclonal antibodies, and stored at a range of laboratory temperatures and lyophilised for distribution and subsequent application.

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Cite as

Di Genova, C., Sampson, A., Scott, S., Cantoni, D., Neto, M., Bentley, E., Mattiuzzo, G., Wright, E., Derveni, M., Auld, B., Ferrara, B., Harrison, D., Said, M., Selim, A., Thompson, E., Thompson, C., Carnell, G. & Temperton, N. 2021, 'Production, titration, neutralisation, storage and lyophilisation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lentiviral pseudotypes', Bio-Protocol Journal, 11(21), article no: e4236. https://doi.org/10.21769/BioProtoc.4236

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DOI: https://doi.org/10.21769/BioProtoc.4236

Repository URI: https://eprints.gla.ac.uk/308334/