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PSEUDOTYPED VIRUSES PERMIT RAPID DETECTION OF NEUTRALIZING ANTIBODIES IN HUMAN AND EQUINE SERUM AGAINST VENEZUELAN EQUINE ENCEPHALITIS VIRUS

ANDREY A. KOLOKOLTSOV, ERYU WANG, TONYA M. COLPITTS, SCOTT C. WEAVER, AND ROBERT A. DAVEY^*

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.

Received January 23, 2006. Accepted for publication May 23, 2006.

Acknowledgments: We thank Ms. Mardelle Susman for editing the manuscript. Thanks to Dr. Robert Tesh for providing characterized mouse serum raised to specific alphaviruses and James Olson and Evelia Quiroz for providing other sera. Technical assistance was provided by Wenli Kang. This work was supported by grants from NIAID to RD and SCW through the Western Regional Center of Excellence for Biodefense and Emerging Infectious Disease Research, NIH Grant number U54 AI057156. Other support to SCW was provided by NIH Grant 48807 and by NIH contract N01-AI25489 and to RD by NIH Grant R56 AI 63513-01.

^* Address correspondence to Robert Davey, Department of Microbiology and Immunology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555. E-mail: radavey{at}utmb.edu

These authors contributed equally to this work.

Authors’ addresses: Andrey A. Kolokoltsov, Tonya M. Colpitts, and Robert A. Davey, 4.138, MRB, Department of Microbiology and Immunology, 301 University Blvd., Galveston, TX 77555. Eryu Wang and Scott C. Weaver, 4.128, Department of Pathology, Kieller Bldg., 301 University Blvd., Galveston, TX 77555.

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