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Studying longitudinal neutralising antibody levels against Equid herpesvirus 1 in experimentally infected horses using a novel pseudotype based assay

Di Genova, Cecilia, Sutton, Gabrielle, Paillot, Romain, Temperton, Nigel J., Pronost, Stéphane, Scott, Simon D. (2023) Studying longitudinal neutralising antibody levels against Equid herpesvirus 1 in experimentally infected horses using a novel pseudotype based assay. Virus Research, 339 . Article Number 199262. ISSN 0168-1702. E-ISSN 1872-7492. (doi:10.1016/j.virusres.2023.199262) (KAR id:104045)


Infection with equid herpesvirus 1 (EHV-1), a DNA virus of the Herpesviridae family represents a significant welfare issue in horses and a great impact on the equine industry. During EHV-1 infection, entry of the virus into different cell types is complex due to the presence of twelve glycoproteins (GPs) on the viral envelope. To investigate virus entry mechanisms, specific combinations of GPs were pseudotyped onto lentiviral vectors. Pseudotyped virus (PV) particles bearing gB, gD, gH and gL were able to transduce several target cell lines (HEK293T/17, RK13, CHO-K1, FHK-Tcl3, MDCK I & II), demonstrating that these four EHV-1 glycoproteins are both essential and sufficient for cell entry. The successful generation of an EHV-1 PV permitted development of a PV neutralisation assay (PVNA). The efficacy of the PVNA was tested by measuring the level of neutralising serum antibodies from EHV-1 experimentally infected horses (n = 52) sampled in a longitudinal manner. The same sera were assessed using a conventional EHV-1 virus neutralisation (VN) assay, exhibiting a strong correlation (r = 0.82) between the two assays. Furthermore, PVs routinely require -80 ◦C for long term storage and a dry ice cold-chain during transport, which can impede dissemination and utilisation in other stakeholder laboratories. Consequently, lyophilisation of EHV-1 PVs was conducted to address this issue. PVs were lyophilised and pellets either reconstituted immediately or stored under various temperature conditions for different time periods. The recovery and functionality of these lyophilised PVs was compared with standard frozen aliquots in titration and neutralisation tests. Results indicated that lyophilisation could be used to stably preserve such complex herpesvirus pseudotypes, even after weeks of storage at room temperature, and that reconstituted EHV-1 PVs could be successfully employed in antibody neutralisation tests.

Item Type: Article
DOI/Identification number: 10.1016/j.virusres.2023.199262
Projects: 215 22924
Uncontrolled keywords: equid herpesvirus 1; lentiviral pseudotype virus; serology; neutralisation assay
Subjects: Q Science
Q Science > QR Microbiology
Q Science > QR Microbiology > QR355 Virology
Divisions: Divisions > Division of Natural Sciences > Medway School of Pharmacy
Funders: University of Kent (
Depositing User: Simon Scott
Date Deposited: 23 Nov 2023 11:34 UTC
Last Modified: 07 Mar 2024 12:27 UTC
Resource URI: (The current URI for this page, for reference purposes)

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