Generation of soluble, disulfide-rich JEV NS1 protein recognizable by anti-NS1 antibodies through a simplified, in vitro refolding approach.

Co-existence of Japanese Encephalitis virus (JEV) with highly homologous antigenic epitopes results in antibody-based serodiagnosis being inaccurate at detecting and distinguishing JEV from other flaviviruses. This often causes misdiagnosis and inefficient treatments of flavivirus infection. Generation of JEV NS1 protein remains a challenge as it is notably expressed in the form of inactive aggregates known as inclusion bodies using bacterial expression systems. This study evaluated two trxB and gor E. coli strains in producing soluble JEV NS1 via a cold-shock expression system. High yield of JEV NS1 inclusion bodies was produced using cold-shocked expression system. Subsequently, a simplified yet successful approach in generating soluble, active JEV NS1 protein through solubilization, purification and in vitro refolding of JEV NS1 protein from inclusion bodies was developed. A step-wise dialysis refolding approach was used to facilitate JEV NS1 refolding. The authenticity of the refolded JEV NS1 was confirmed by specific antibody binding on indirect ELISA commercial anti-NS1 antibodies which showed that the refolded JEV NS1 was highly immunoreactive. This presented approach is cost-effective, and negates the need for mammalian or insect cell expression systems in order to synthesize this JEV NS1 protein of important diagnostic and therapeutic relevance in Japanese Encephalitis disease.

Flavivirus infection-A review of immunopathogenesis, immunological response, and immunodiagnosis.

Flaviviruses are group of single stranded RNA viruses that cause severe endemic infection and epidemics on a global scale. It presents a significant health impact worldwide and the viruses have the potential to emerge and outbreak in a non-endemic geographical region. Effective vaccines for prophylaxis are only available for several flaviviruses such as Yellow Fever virus, Tick-borne Encephalitis Virus, Dengue Virus and Japanese Encephalitis Virus and there is no antiflaviviral agent being marketed. This review discusses the flavivirus genome, replication cycle, epidemiology, clinical presentation and pathogenesis upon infection. Effective humoral response is critical to confer protective immunity against flaviviruses. Hence, we have also highlighted the immune responses elicited upon infection, various diagnostic facilities available for flaviviral disease and monoclonal antibodies available to date against flavivirus infection.