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

Chumbe, Ana, Urbanowicz, Richard A ORCID: 0000-0002-2461-4993, Sliepen, Kwinten, Koekkoek, Sylvie M, Molenkamp, Richard, Tarr, Alexander W, Ball, Jonathan K, Schinkel, Janke and van Gils, Marit J (2022) Optimization of the pseudoparticle system for standardized assessments of neutralizing antibodies against hepatitis C virus. The Journal of general virology, 103 (11).

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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.

Item Type:	Article
Uncontrolled Keywords:	antibodies, HCV, neutralization, optimization, pseudoparticles, standardization
Divisions:	Faculty of Health and Life Sciences Faculty of Health and Life Sciences > Institute of Infection, Veterinary and Ecological Sciences
Depositing User:	Symplectic Admin
Date Deposited:	22 Dec 2022 11:47
Last Modified:	06 Sep 2023 17:50
DOI:	10.1099/jgv.0.001801
Open Access URL:	https://www.microbiologyresearch.org/content/journ...
Related URLs:	Author Publisher
URI:	https://livrepository.liverpool.ac.uk/id/eprint/3166755