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Optimization of the pseudoparticle system for standardized assessments of neutralizing antibodies against hepatitis C virus

Chumbe, Ana; Urbanowicz, Richard A.; Sliepen, Kwinten; Koekkoek, Sylvie M.; Molenkamp, Richard; Tarr, Alexander W.; Ball, Jonathan K.; Schinkel, Janke; van Gils, Marit J.

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Authors

Ana Chumbe

Richard A. Urbanowicz

Kwinten Sliepen

Sylvie M. Koekkoek

Richard Molenkamp

JONATHAN BALL jonathan.ball@nottingham.ac.uk
Professor of Molecular Virology

Janke Schinkel

Marit J. van Gils



Abstract

A better understanding of the antibody response during natural infection and the effect on disease progression and reinfection is necessary for the development of a protective hepatitis C virus (HCV) vaccine. The HCV pseudoparticle (HCVpp) system enables the study of viral entry and inhibition by antibody neutralization. A robust and comparable neutralization assay is crucial for the development and evaluation of experimental vaccines.With the aim of optimizing the HCVpp-murine leukaemia virus (MLV) system, we tested the neutralization of HCVpp-harbouring E1E2 from 21 HCV isolates representing 6 different genotypes by several monoclonal antibodies (mAbs). HCVpps are generated by expressing functional envelope glycoproteins (E1E2) onto pseudoparticles derived from env-deleted MLV. Adjustments of E1E2, gag-pol and luciferase plasmid ratios resulted in increased yields for most HCVpps and recovery of one non-infectious HCVpp. We simplified and improved the protocol to achieve higher signal/noise ratios and minimized the amount of HCVpps and mAbs needed for the detection of neutralization. Using our optimized protocol, we demonstrated comparable results to previously reported data with both diluted and freeze-thawed HCVpps.In conclusion, we successfully established a simplified and reproducible HCVpp neutralization protocol for studying a wide range of HCV variants. This simplified protocol provides highly consistent results and could be easily adopted by others to evaluate precious biological material. This will contribute to a better understanding of the antibody response during natural infection and help evaluate experimental HCV vaccines.

Citation

Chumbe, A., Urbanowicz, R. A., Sliepen, K., Koekkoek, S. M., Molenkamp, R., Tarr, A. W., …van Gils, M. J. (2022). Optimization of the pseudoparticle system for standardized assessments of neutralizing antibodies against hepatitis C virus. Journal of General Virology, 103(11), Article 001801. https://doi.org/10.1099/jgv.0.001801

Journal Article Type Article
Acceptance Date Oct 3, 2022
Online Publication Date Nov 18, 2022
Publication Date 2022
Deposit Date Nov 24, 2022
Publicly Available Date Mar 29, 2024
Journal Journal of General Virology
Print ISSN 0022-1317
Electronic ISSN 1465-2099
Publisher Microbiology Society
Peer Reviewed Peer Reviewed
Volume 103
Issue 11
Article Number 001801
DOI https://doi.org/10.1099/jgv.0.001801
Keywords Virology
Public URL https://nottingham-repository.worktribe.com/output/13754481
Publisher URL https://www.microbiologyresearch.org/content/journal/jgv/10.1099/jgv.0.001801