Estabalishment And Preliminary Application Of Hybridoma Cell Lines Secreting Monoclonal Antibodies Against Pseudorabies Virus

BALB/C mice were immunized with the purified PrV of MIN A isolate,mouse spleen cell were fused with myelomas NSo.Culture supernatants of hybridomas were screened by indirect ELISA and immobilization ELISA,positive colonies were cloned three times by limited dilution.Only one cell line named OHe which could secrete McAb stably was obtained.The isotype of the McAb was IgGi,the average number of chromossme of the hybridoma cell line was 97 respectively.The ELISA titer of cell supernate and ascite were l:1024and l:204800.The McAb did not cross react with PRRSV, HCV, PPV,demonstrated good specifity.The OHecell line steadly secreted McAb after subcultured twenty-one times .Antigen-capture-ELISA for detecting PrV was developed by using OH6 strain monoclonal antibody. The optimal coating concentration of McAb was 2.04ug/ml,working concentration of rabbit-anti-PrV hyperimmune serum was l:8000.The sensitivity of the McAb-AC-ELISA was 5.5ng/ml.The assay had no cross reaction with common pathogenic microorganisms,and the positive coincidence rate with virus isolation assay was high.The results demonstrated that positive rates in tonsiK brain and lung were higher than that in other tissues by detecting PrV antigen in tonsiK liver> spleen. lung> kindey and brain of pigs infected by PrV. Establishment of a McAb-AC-ELISAprovided a simple , fast, sensitive and universally used method for the diagnosis ofPrV.