Antigenic Epitopes Identification Of Duck Tembusu Virus Envelope Protein

An antigenic epitope, also known as antigenic determinant, is the part of an antigen thatis recognized by the immune system, specifically by antibodies, B cells, or T cells. Antigenicepitope mapping not only analysis the structure and function of protein antigens, but alsoprepare a safe and effective vaccines and diagnostic reagents.Duck tembusu disease is an important insect-borne infectious disease, and the affectedducks showed a severe drop in egg production and paralysis. The disease has caused seriouseconomy in aquaculture in south of China since2010. The researches of DTMUV have madeimportant progresses. However, there was not any literature about the angentic epitopemapping on DTMUV envelope protein. So epitope mapping is an assistant for designing aserological diagnosis method, besides, it provided the theory foundation for vaccines designof envelope protein.The envelope protein gene of duck Tembusu virus （DTMUV） was amplified by RT-PCRfrom DTMUV YY5-infected DF-1cells and cloned into pMBP-c prokaryotic expressionvector, the result shows the MBP-E fusion protein was expressed in soluble form and hasgood immunogenicity and reactionogenicity. Balb/c mice were immunized by using theimmunogen of purified recombinant fusion protein MBP-E, two hybridoma cell lines thatstably secreted monoclonal antibodies against envelop protein after cell fusion andsubcloning, which named BD10and AE₄.The Western blot and IFA results indicated that theepitopes recognized by BD₁₀is lineal determinant, and AE4is conformentional determinant.According to the antigenic epitope of this antigenic dominant region, a set of15partiallyoverlapping fragments spanning the fragment were fused with MBP and expressed.WithWestern blot and ELISA analysis, a B-cell linear antigenic epitopes were identified.Thebalb/c mince were immunized with the purified recombination protein MBP-EP385,the result shows that the MBP-EP385has good immunogenicity and reactionogenicity. The B-celllinear epitope was id all, the minimum function epitope region is EP385(₃₈₅LVGSGKGQ₃₉₃).