Quantification of SARS-CoV-2 neutralizing antibody by wild-type plaque reduction neutralization, microneutralization and pseudotyped virus neutralization assays

Kevin Bewley*, Naomi Coombes, Luc Gagnon, Lorna McInroy, Natalie Baker, Imam Shaik, Julien R. St-Jean, Natalie St-Amant, Karen Buttigieg, Holly Humphries, Kerry J. Godwin, Emily Brunt, Lauren Allen, Stephanie Leung, Phillip J. Brown, Elizabeth J. Penn, Kelly Thomas, Greg Kulnis, Bassam Hallis, Miles CarrollSimon Funnell, Sue Charlton

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Citations (Scopus)

Abstract

Virus neutralization assays measure neutralizing antibodies in serum and plasma, and the plaque reduction neutralization test (PRNT) is considered the gold standard for measuring levels of these antibodies for many viral diseases. We have developed procedures for the standard PRNT, microneutralization assay (MNA) and pseudotyped virus neutralization assay (PNA) for severe acute respiratory syndrome coronavirus 2. The MNA offers advantages over the PRNT by reducing assay time, allowing increased throughput and reducing operator workload while remaining dependent upon the use of wild-type virus. This ensures that all severe acute respiratory syndrome coronavirus 2 antigens are present, but Biosafety Level 3 facilities are required. In addition to the advantages of MNA, PNA can be performed with lower biocontainment (Biosafety Level 2 facilities) and allows for further increases in throughput. For each new vaccine, it is critical to ensure good correlation of the neutralizing activity measured using PNA against the PRNT or MNA. These assays have been used in the development and licensure of the ChAdOx1 nCoV-19 (AstraZeneca; Oxford University) and Ad26.COV2.S (Janssen) coronavirus disease 2019 vaccines and are critical for demonstrating bioequivalence of future vaccines.

Original languageEnglish
Pages (from-to)3114-3140
Number of pages35
JournalNature Protocols
Volume16
Issue number6
Early online date23 Apr 2021
DOIs
Publication statusPublished - Jun 2021

Bibliographical note

Funding Information:
The authors thank C. Cardonnel, field application scientist at Molecular Devices, for her assistance in developing the SoftMax Pro formulae and protocols for the microneutralization assay. The PNA elements of this work were supported by the Global Health Vaccine Accelerator Platforms (GH-VAP) and the Coronavirus Immunotherapy Consortium (CoVIC) funded by BMGF.

Publisher Copyright:
© 2021, The Author(s), under exclusive licence to Springer Nature Limited.

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