Japanese encephalitis virus (JEV), a mosquito-borne neurotropic flavivirus, causes severe diseases of the central nervous system in humans. Gold nanoparticle increases the sensitivity for in vivo and in vitro detection, being widely used in biomedical applications. We investigated the sensitivity of gold nanopaticles conjugated to JEV-specific Envelope protein domain III compared with the ELISA assay for diagnosis of JEV infection. For the preparation of JEV-specific polyclonal and monoclonal antibody, BALB/c mice were immunized with recombinant JEV envelope domain III (EDIII) protein with adjuvant every two week five times. JEV-specific mAb was identified from hybridoma cells that are fused by spleen cells from immunized mice and myeloma cells. JEV-specific mAb had a binding specificity to recombinant JEV EDIII protein and JEV virions using Western blotting and ELISA. Polyclonal antibodies were made from New Zealand White rabbit immunized with JEV EDIII protein. In addition, JEV-specific mAb showed the neutralizing ability against JEV. Subsequently, ED3-linked gold nanoparticles were demonstrated higher sensitivity of JEV detection than conventional ELISA. Meanwhile, ED3-linked gold nanoparticles were applied on the ELISA assay in JEV infected serum. Polyclonal and monoclonal antibodies were also conjugated with gold nanoparticles for detection of JEV in future. Therefore, gold nanoparticles conjugated to virus-specific protein will be more fast and sensitive on viral detection.
