Volume 75, Issue 4
  • ISSN: 0002-9637
  • E-ISSN: 1476-1645


Virus envelope proteins are the primary targets of neutralizing antibody responses. The epitopes recognized differ sufficiently between virus subtypes and species to distinguish viruses and provide an important basis for disease diagnosis. Venezuelan equine encephalitis virus (VEEV) causes acute febrile illness in humans and has high mortality in equines. The most specific detection methods for serum antibodies use live virus in neutralization assays or in blocking enzyme linked immunosorbent assays. However, work with Venezuelan equine encephalitis virus requires biosafety level 3 containment and select agent security in the United States. We report two new assays for detection of Venezuelan equine encephalitis virus neutralizing antibody responses, based on virus pseudotypes. The first provides detection by marker gene expression after 20 hours and is particularly suited for high-throughput screening; the second uses a new, rapid virus entry assay to give readouts within 1 hour. Both assays are safe, sensitive, and in general recapitulate neutralizing antibody titers obtained by conventional plaque reduction assays. Each is suitable as a rapid primary screen for detection of neutralizing antibodies against Venezuelan equine encephalitis virus.


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  1. Johnson KM, Martin DH, 1974. Venezuelan equine encephalitis. Adv Vet Sci Comp Med 18 : 79–116. [Google Scholar]
  2. Fact sheet: Arboviral Encephalitis: CDC.
  3. Canonico PG, Kende M, Luscri BJ, Huggins JW, 1984. In-vivo activity of antivirals against exotic RNA viral infections. J Antimicrob Chemother 14 (Suppl A): 27–41. [Google Scholar]
  4. Kinney RM, Chang GJ, Tsuchiya KR, Sneider JM, Roehrig JT, Woodward TM, Trent DW, 1993. Attenuation of Venezuelan equine encephalitis virus strain TC-83 is encoded by the 5′-noncoding region and the E2 envelope glycoprotein. J Virol 67 : 1269–1277. [Google Scholar]
  5. Roehrig JT, Mathews JH, 1985. The neutralization site on the E2 glycoprotein of Venezuelan equine encephalomyelitis (TC-83) virus is composed of multiple conformationally stable epitopes. Virology 142 : 347–356. [Google Scholar]
  6. Kolokoltsov AA, Weaver SC, Davey RA, 2005. Efficient functional pseudotyping of oncoretroviral and lentiviral vectors by Venezuelan equine encephalitis virus envelope proteins. J Virol 79 : 756–763. [Google Scholar]
  7. Kolokoltsov AA, Davey RA, 2004. Rapid and sensitive detection of retrovirus entry by using a novel luciferase-based content-mixing assay. J Virol 78 : 5124–5132. [Google Scholar]
  8. Berge TO, Banks IS, Tigertt WD, 1961. Attenuation of Venezuelan equine encephalomyelitis virus by in vitro cultivation in guinea pig heart cells. Am J Hyg 73 : 209–218. [Google Scholar]
  9. Weaver SC, Salas R, Rico-Hesse R, Ludwig GV, Oberste MS, Boshell J, Tesh RB, 1996. Re-emergence of epidemic Venezuelan equine encephalomyelitis in South America. VEE Study Group. Lancet 348 : 436–440. [Google Scholar]
  10. Wang E, Paessler S, Aguilar PV, Smith DR, Coffey LL, Kang W, Pfeffer M, Olson J, Blair PJ, Guevara C, Estrada-Franco J, Weaver SC, 2005. A novel, rapid assay for detection and differentiation of serotype-specific antibodies to Venezuelan equine encephalitis complex alphaviruses. Am J Trop Med Hyg 72 : 805–810. [Google Scholar]
  11. Chen C, Okayama H, 1987. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol 7 : 2745–2752. [Google Scholar]
  12. Kolokoltsov AA, Fleming EH, Davey RA, 2005. Venezuelan equine encephalitis virus entry mechanism requires late endosome formation and resists cell membrane cholesterol depletion. Virology.
  13. Beaty BJ, Calisher CH, Shope RE, 1989. Arboviruses. Schmidt NJ, Emmons RW, eds. Diagnostic Procedures for Viral, Rickettsial and Chlamydial Infections. Washington DC: American Public Health Association, 797–855.
  14. National Institutes of Health, 2002. Guidelines for Research Involving Recombinant DNA Molecules: NIH.
  15. Kinney RM, Trent DW, 1982. Conservation of tryptic peptides in the structural proteins of viruses in the Venezuelan equine encephalitis complex. Virology 121 : 345–362. [Google Scholar]
  16. Roehrig JT, Day JW, Kinney RM, 1982. Antigenic analysis of the surface glycoproteins of a Venezuelan equine encephalomyelitis virus (TC-83) using monoclonal antibodies. Virology 118 : 269–278. [Google Scholar]
  17. Hunt AR, Johnson AJ, Roehrig JT, 1990. Synthetic peptides of Venezuelan equine encephalomyelitis virus E2 glycoprotein. I. Immunogenic analysis and identification of a protective peptide. Virology 179 : 701–711. [Google Scholar]
  18. Sandrin V, Muriaux D, Darlix JL, Cosset FL, 2004. Intracellular trafficking of Gag and Env proteins and their interactions modulate pseudotyping of retroviruses. J Virol 78 : 7153–7164. [Google Scholar]
  19. Zavada J, 1982. The pseudotypic paradox. J Gen Virol 63 : 15–24. [Google Scholar]
  20. Sanders DA, 2002. No false start for novel pseudotyped vectors. Curr Opin Biotechnol 13 : 437–442. [Google Scholar]
  21. Bartosch B, Bukh J, Meunier JC, Granier C, Engle RE, Blackwelder WC, Emerson SU, Cosset FL, Purcell RH, 2003. In vitro assay for neutralizing antibody to hepatitis C virus: evidence for broadly conserved neutralization epitopes. Proc Natl Acad Sci USA 100 : 14199–14204. [Google Scholar]
  22. Han DP, Kim HG, Kim YB, Poon LL, Cho MW, 2004. Development of a safe neutralization assay for SARS-CoV and characterization of S-glycoprotein. Virology 326 : 140–149. [Google Scholar]
  23. Rother RP, Fodor WL, Springhorn JP, Birks CW, Setter E, Sandrin MS, Squinto SP, Rollins SA, 1995. A novel mechanism of retrovirus inactivation in human serum mediated by anti-alpha-galactosyl natural antibody. J Exp Med 182 : 1345–1355. [Google Scholar]
  24. Young NA, Johnson KM, 1969. Antigenic variants of Venezuelan equine encephalitis virus: their geographic distribution and epidemiologic significance. Am J Epidemiol 89 : 286–307. [Google Scholar]
  25. Weaver SC, Ferro C, Barrera R, Boshell J, Navarro JC, 2004. Venezuelan equine encephalitis. Annu Rev Entomol 49 : 141–174. [Google Scholar]

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  • Received : 23 Jan 2006
  • Accepted : 23 May 2006

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