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BioMed Research International
Volume 2015, Article ID 678084, 13 pages
Research Article

A High-Performance Multiplex Immunoassay for Serodiagnosis of Flavivirus-Associated Neurological Diseases in Horses

1UMR 1161 of Virology, ANSES, INRA, ENVA, ANSES Animal Health Laboratory, EU-RL on Equine Diseases, UPE, 94701 Maisons-Alfort, France
2UMR PIMIT (I2T Team), INSERM U1187, CNRS 9192, IRD 249, Technology Platform CYROI, University of Reunion, 97491 Saint-Clotilde, Réunion
3Department of Infections and Epidemiology, Institut Pasteur, 75724 Paris, France
4Viral Zoonoses, Emerging and Vector-Borne Infections Group, Institute of Virology, University of Veterinary Medicine Vienna, 1210 Vienna, Austria
5Department of Microbiology and Immunology, College of Medicine and Health Sciences, Sultan Qaboos University, 123 Al-Khoudh, Oman
6INRA UE 1277, Plate-Forme d’Infectiologie Expérimentale, 37380 Nouzilly, France
7Equine Research Institute, Japan Racing Association, Tochigi 329-0412, Japan

Received 11 March 2015; Accepted 12 May 2015

Academic Editor: Sulagna Das

Copyright © 2015 Cécile Beck et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


West Nile virus (WNV), Japanese encephalitis virus (JEV), and tick-borne encephalitis virus (TBEV) are flaviviruses responsible for severe neuroinvasive infections in humans and horses. The confirmation of flavivirus infections is mostly based on rapid serological tests such as enzyme-linked immunosorbent assays (ELISAs). These tests suffer from poor specificity, mainly due to antigenic cross-reactivity among flavivirus members. Robust diagnosis therefore needs to be validated through virus neutralisation tests (VNTs) which are time-consuming and require BSL3 facilities. The flavivirus envelope (E) glycoprotein ectodomain is composed of three domains (D) named DI, DII, and DIII, with EDIII containing virus-specific epitopes. In order to improve the serological differentiation of flavivirus infections, the recombinant soluble ectodomain of WNV E (WNV.sE) and EDIIIs (rEDIIIs) of WNV, JEV, and TBEV were synthesised using the Drosophila S2 expression system. Purified antigens were covalently bonded to fluorescent beads. The microspheres coupled to WNV.sE or rEDIIIs were assayed with about 300 equine immune sera from natural and experimental flavivirus infections and 172 nonimmune equine sera as negative controls. rEDIII-coupled microspheres captured specific antibodies against WNV, TBEV, or JEV in positive horse sera. This innovative multiplex immunoassay is a powerful alternative to ELISAs and VNTs for veterinary diagnosis of flavivirus-related diseases.