Developing Pseudovirus-Based Neutralization Assay against Omicron-Included SARS-CoV-2 Variants

Abstract

The global spread of SARS-CoV-2 and its variants poses a serious threat to human health worldwide. Recently, the emergence of Omicron has presented a new challenge to the prevention and control of the COVID-19 pandemic. A convenient and reliable in vitro neutralization assay is an important method for validating the efficiency of antibodies, vaccines, and other potential drugs. Here, we established an effective assay based on a pseudovirus carrying a full-length spike (S) protein of SARS-CoV-2 variants in the HIV-1 backbone, with a luciferase reporter gene inserted into the non-replicate pseudovirus genome. The key parameters for packaging the pseudovirus were optimized, including the ratio of the S protein expression plasmids to the HIV backbone plasmids and the collection time for the Alpha, Beta, Gamma, Kappa, and Omicron pseudovirus particles. The pseudovirus neutralization assay was validated using several approved or developed monoclonal antibodies, underscoring that Omicron can escape some neutralizing antibodies, such as REGN10987 and REGN10933, while S309 and ADG-2 still function with reduced neutralization capability. The neutralizing capacity of convalescent plasma from COVID-19 convalescent patients in Wuhan was tested against these pseudoviruses, revealing the immune evasion of Omicron. Our work established a practical pseudovirus-based neutralization assay for SARS-CoV-2 variants, which can be conducted safely under biosafety level-2 (BSL-2) conditions, and this assay will be a promising tool for studying and characterizing vaccines and therapeutic candidates against Omicron-included SARS-CoV-2 variants.

SARS-CoV-2 Omicron subvariants Spike recognition and neutralization elicited after the third dose of mRNA vaccine

“…Finally, while we did not observe major differences in infectivity with our pseudoviral particles, it is possible that differences in infectivity and replication exist when using authentic live viruses. For this reason we only report on plasma neutralization profiles, which were shown to be similar between pseudoviral particles and authentic viruses and have been largely used by the field to inform on neutralizing responses elicited by natural infection and vaccination [42][43][44][45][46]…”

Presence of neutralizing SARS‐CoV‐2 antibodies in asymptomatic population of N'Djamena, Chad

“…Alternative SARS‐CoV‐2 pseudovirus technology uses lentiviral particles based on human immunodeficiency virus, retroviral particles based on murine leukemia virus, or systems based on vesicular stomatitis virus (VSV), have been developed to avoid the culture of live SARS‐CoV‐2. The results from such pseudovirus neutralization assays correlate well with the results of measurements using authentic viruses 12–14 . Although alternative BSL2 protocols using pseudotyped have been developed, these methods remain in the research area 15 …”

“…Since the beginning of the Covid‐19 pandemic, several highly transmissible variants of SARS‐CoV‐2 have emerged 16,17 . Some mutations in the SARS‐CoV‐2 spike protein (S) can alter antigenic properties, allowing them to be more transmissible, virulent, pathogenic or evade immunity induced by previous infection 12 . In this situation, it would be important to consider these mutations in the production process of pseudoviruses to assess the efficacy of NAb production in vaccines, patients, and asymptomatic individuals.…”

Gold nanoparticle-based immunochromatographic assay for the rapid detection of the SARS-CoV-2 Omicron variant

“…8,9 Pseudo virus neutralization tests are costly and difficult to perform and are not suitable for use in pandemic disease testing. [10][11][12] Antibody-based assays have the potential to be used to improve diagnostic efficiency, and currently, the US Food and Drug Administration has approved a variety of rapid lateral flow immunoassays based on gold nanoparticles (GNPs), 13 fluorescent microspheres 14 and quantum dots 15,16 for the diagnosis of SARS-CoV-2 infection. However, previous tests have been developed based on wild-type strains.…”