Generation of VSV pseudotypes using recombinant ΔG-VSV for studies on virus entry, identification of entry inhibitors, and immune responses to vaccines.
Vesicular stomatitis virus (VSV) is a prototypic enveloped animal virus that has been used extensively to study virus entry, replication and assembly due to its broad host range and robust replication properties in a wide variety of mammalian and insect cells. Studies on VSV assembly led to the creation of a recombinant VSV in which the glycoprotein (G) gene was deleted. This recombinant (rVSV-ΔG) has been used to produce VSV pseudotypes containing the envelope glycoproteins of heterologous viruses, including viruses that require high-level biocontainment; however, because the infectivity of rVSV-ΔG pseudotypes is restricted to a single round of replication the analysis can be performed using biosafety level 2 (BSL-2) containment. As such, rVSV-ΔG pseudotypes have facilitated the analysis of virus entry for numerous viral pathogens without the need for specialized containment facilities. The pseudotypes also provide a robust platform to screen libraries for entry inhibitors and to evaluate the neutralizing antibody responses following vaccination. This manuscript describes methods to produce and titer rVSV-ΔG pseudotypes. Procedures to generate rVSV-ΔG stocks and to quantify virus infectivity are also described. These protocols should allow any laboratory knowledgeable in general virological and cell culture techniques to produce successfully replication-restricted rVSV-ΔG pseudotypes for subsequent analysis.
Whitt
Michael A
MA
Department of Molecular Sciences, 858 Madison Ave., The University of Tennessee Health Science Center, Memphis, TN 38163, USA. mwhitt@uthsc.edu
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D017353:Gene Deletion; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D018116:Vesiculovirus / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000502:physiology*; D019562:Viral Load; D014765:Viral Vaccines / Q000235:genetics / Q000276:immunology*; D014776:Virus Cultivation; D053586:Virus Internalization / Q000187:drug effects*
D000998:Antiviral Agents; C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D014765:Viral Vaccines