Pseudovirus-Based Assays for the Measurement of Antibody-Mediated Neutralization of SARS-CoV-2

Corey Balinsky*, Vihasi Jani, Peifang Sun, Maya Williams, Gabriel Defang, Kevin R. Porter

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

1 Scopus citations


SARS-CoV-2 has emerged as a significant cause of morbidity and mortality worldwide. Virus neutralization assays are critical for the development and evaluation of vaccines and immunotherapeutics, as well as for conducting basic research into the immune response, spread, and pathogenesis of this disease. However, neutralization assays traditionally require the use of infectious virus which must be carefully handled in a BSL-3 setting, thus complicating the assay and restricting its use to labs with access to BSL-3 facilities. Pseudovirus-based assays are an alternative to the use of infectious virus. SARS-CoV-2 pseudovirus contains only the spike structural protein, and infection results in a single round of replication, thus allowing for the assay to be run safely under BSL-2 conditions. In this chapter, we describe protocols and considerations for the production and titration of lentivirus-based SARS-CoV-2 pseudovirus, as well as for running and analysis of FACS-based pseudovirus neutralization assays.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Number of pages18
StatePublished - 2022
Externally publishedYes

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029


  • Antibody
  • COVID-19
  • Lentivirus
  • Neutralization
  • Protocols
  • Pseudovirus (PV)
  • SARS-CoV-2
  • Spike


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