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. 2020 Apr 10;12(4):429. doi: 10.3390/v12040429

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© 2020 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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Construction of stable cell lines expressing bovine herpesvirus 1 (BoHV-1), pseudorabies virus (PRV), or herpes simplex 1 (HSV-1) glycoprotein B (gB), and characterization of size exclusion chromatography (SEC)-isolated extracellular vesicles (EVs). (A) The list of constructed cell lines expressing individual viral gB homologs. Human melanoma Mel JuSo (MJS) and Madin–Darby bovine kidney (MDBK) produce endogenous major histocompatibility (MHC) class II. The Human Jurkat T cell line and swine kidney SK6 cells are MHC II-negative. (B–E) Immunoblotting detection of EVs markers Alix and flotillin-2 (flot-2) in cell lysates (cl) of constructed cell lines or SEC-isolated EVs. Calnexin (CNX) and mitochondrial Tom 40 were used as non-EVs control proteins. MJSpuro, MDBKpuro, SK6puro, or untransduced Jurkat cells (Ø) were used as negative controls. (F) Representative transmission electron microscopy images of EVs preparations from gB-expressing species-specific cells; scale bar 100 nm.