Generation Of Monoclonal Antibodies And Epitope Identification For VP22 Of Duck Enteritis Virus

Duck viral enteritis (DVE), also known as duck plague (DP), is an acute, heat and haemorrhagic contagious viral disease that naturally affecting birds of the order Anseriformes (ducks, geese and swans). DVE was caused by duck enteritis virus (DEV). This disease was epidemic in many countries and regions and caused economic losses. Antigenic epitope information of DEV is not only useful for investigating the relationship between antigenic structure and function of the DEV, but also significant for diagnosis of DEV infection and developing safe and effective multi-epitope vaccine against the disease.The E.coli BL21 (DE3) contained the recombinant plasmid pET-30a-UL49 was induced by IPTG and the recombinant protein was analyzed by SDS-PAGE. The recombinant protein with a molecular weight of 36 ku could be detected by Western blot using the antiserum against DEV. Monoclonal antibodies (MAbs) against VP22 protein of DEV were developed using recombinant protein VP22 as antigen. Four mAbs against VP22 were obtained following screening by indirect enzyme-linked immunosorbent assay (ELISA), Western blot and indirect immunofluorescence assay. Four MAbs, designated as 2B10, 2G1, 3F10 and 3G2 respectively, could recognize the VP22 protein in DEV-infected chicken embryo fibroblasts. This study paved the way for future study of biological function and antigenic epitope of VP22 protein.The pepscan procedure was used to identify B-cell epitopes by using 20 recombinant fragments expressed as GST-fusion proteins. The fragments were analyzed by Western blot and enzyme-linked immunosorbent assay (ELISA) with representative IgG2b MAb, 2B10, which exhibited a strong immunoreaction to both recombinant VP22 and native viral protein DEV. Results indicated that the motif 200ELSGQTP206 is the minimal epitope recognized by MAb 2B10, and the epitope could also be recognized by DEV-postive serum from ducks. These findings not only suggest that VP22 specific MAbs may be used as the diagnostic assays for DEV, but also provide a basis development of epitope-based marker vaccine against DEV.