| Spring Viremia of Carp Virus is the pathogen of infectious hemorrhagic swimbladder inflammation in common carp (Cyprinus Carpio) and the mortality of juvenile carp during spring outbreaks can be30-70%. Thus, it has been listed as notifiable in the International Aquatic Animal Health Code of the Office International des Epizooties (OIE,2006). The initial outbreak of Spring Viremia of Carp (SVC) was in European countries. Recently, it has been reported in the Middle East, USA, China, Canada and Brazil.The SVCV glycoprotein gene segment was amplified by PCR and cloned into the prokaryotic expression vector pGEX-KG. The SVCV-g-KG fusion protein(66KDa) was expressed highly under induction of IPTG in the E.coli BL21, and this fusion protein was mainly expressed in the insoluble composition.Two clones of monoclonal antibodies (MAbs1H11and4B8) against G protein were generated by fusion of mouse myeloma cell line SP2/0and spleen lymphocytes from part of G protein (3094-4170bp) immunized mice. The results of ELISA (Enzyme linked immunosorbent assay), IFA (Indirect immunofluorescent assay) and Western-blot assay further demonstrated the characterizations of the two MAbs. Both1H11and4B8were specific to SVCV G protein.Then, a set of synthesized peptides were coated as antigen in ELISA for further epitope mapping. Finally, the amino acids273-287(DGTLVSGHRPGLDLI) were identified as the epitope of G protein.The G protein-specific polyclonal antibody was prepared using the Japanese white rabbits, which were immuned with the purified recombinant G protein. The ELISA titer of the polyclonal antibody was1:256000. Western blot and indirect immunofluorescent assay identified that the polyclonal antibody has high specificity. Neutralization test of polyclonal antibody showed that it could stop the virus from infecting the cell efficiently. These specific MAbs and PAbs can be useful tools for developing rapid detection methods for SVCVand research of the pathogenic mechanism of SVCV. |
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