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Preparation of monoclonal antibodies against duck enteritis virus
WANG Honghai1, SU Jingliang1, CAO
Zhen2, TIAN Kegong2
(1. College of Veterinary Medicine，China Agricultural University，Beijing 100094,China;
2. National Veterinary Diagnostic Center, Ministry of Agriculture，Beijing 100094,China) |
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Abstract: Duck enteritis virus(DEV)was propagated in duck embryo fibroblasts(DEF) and purified by ultracentrifugation. Four BALB/c mice were immunized subcutaneously with purifiedDEV antigen three times at 10 days interval. The immunized spleen cellsof the mice were fused with the SP2/0 myeloma cells. An indirect ELISA was developed to detect DEV antigenspecific monoclonal antibodies(McAb) in cell culture supernatants. Two hybridoma cell lines, named 1B6and 2G8 ,secreting McAb against DEV were prepared after three times of limitingdilution cloning. The immunoglobulin subclass of 1B6and 2G8 was IgM and IgG1 respectively. The ELISA titer of ascites fluid collected from mice inoculated with either of the two hyridomas were all above
1∶107. A McAb based indirect immunoflurescent antibody assay was established to detect DEV antigen in DEF using purified 2G8ascites immunoglobulin. The optimal working dilution of the McAb was 1∶100. The viral antigen could be detected out in 6 hours postinoculation. The immunofluorescence was most significant with relativelyintact cell morphorology.
Key words: duck enteritis virus; monoclonal antibody; preparation |
