Deletion of the SARS-CoV-2 Spike Cytoplasmic Tail Increases Infectivity in Pseudovirus Neutralization Assays

• Published in: Journal of virology Vol. 95; no. 11
• Main Authors: Yu, Jingyou, Li, Zhenfeng, He, Xuan, Gebre, Makda S, Bondzie, Esther A, Wan, Huahua, Jacob-Dolan, Catherine, Martinez, David R, Nkolola, Joseph P, Baric, Ralph S, Barouch, Dan H
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 10.05.2021
• Subjects: Vaccines and Antiviral Agents; neutralization assay; neutralizing antibodies; SARS-CoV-2; antibody; lentiviral vector; spike
• ISSN: 0022-538X; 1098-5514
• DOI: 10.1128/jvi.00044-21

Pseudotyped viruses are valuable tools for studying virulent or lethal viral pathogens that need to be handled in biosafety level 3 (BSL-3) or higher facilities. With the explosive spread of the coronavirus disease 2019 (COVID-19) pandemic, the establishment of a BSL-2 adapted SARS-CoV-2 pseudovirus neutralization assay is needed to facilitate the development of countermeasures. Here we describe an approach to generate a single-round lentiviral vector-based SARS-CoV-2 pseudovirus, which produced a signal more than 2 logs above background. Specifically, a SARS-CoV-2 spike variant with a cytoplasmic tail deletion of 13 amino acids, termed SΔCT13, conferred enhanced spike incorporation into pseudovirions and increased viral entry into cells as compared with full-length spike (S). We further compared S and SΔCT13 in terms of their sensitivity to vaccine sera, purified convalescent IgG, hACE2-mIgG, and the virus entry inhibitor BafA1. We developed a SΔCT13-based pseudovirus neutralization assay and defined key assay characteristics, including linearity, limit of detection, and intra- and intermediate-assay precision. Our data demonstrate that the SΔCT13-based pseudovirus shows enhanced infectivity in target cells, which will facilitate the assessment of humoral immunity to SARS-CoV-2 infection, antibody therapeutics, and vaccination. This pseudovirus neutralization assay can also be readily adapted to SARS-CoV-2 variants that emerge. SARS-CoV-2 is the etiologic agent of the COVID-19 pandemic. The development of a high throughput pseudovirus neutralization assay is critical for the development of vaccines and immune-based therapeutics. In this study, we show that deletion of the cytoplasmic tail of the SARS-CoV-2 spike leads to pseudoviruses with enhanced infectivity. This SΔCT13-based pseudovirus neutralization assay should be broadly useful for the field.