Development Of Monoclonal Antibodies To UL13Protein Of Marek’s Disease Virus And Its Preliminary Application

1Development and Characterization of Monoclonal Antibody to UL13protein of Marek’s disease virusOur laboratory find out in Genbank published on Marek’s disease virus UL13gene sequences of different strains, Found that the gene sequences of various strains is consistent. Using DNAstar analysis software translated CVI988virus UL13protein sequence into the corresponding protein. Re-use analysis software to analyze the protein, obtain the sites of the high hydrophilic areas,high expression of antigen. Choose the high antigenicity and high hydrophilic peptides, synthetic peptides.Use the high antigenicity,high hydrophilic small peptide as immunogen for immunization of mice. Intraperitoneal immunization8weeks Balb/c mice. First:Mixture Complete adjuvant and peptide, intraperitoneal immunization0.2-0.3ml, containing peptides20ug/one Mouse.The second time in two weeks:incomplete adjuvant and peptide mixture, intraperitoneal immunization0.2-0.3ml, containing peptides50u/one Mouse. The third time in two weeks: incomplete adjuvant and peptide mixture, intraperitoneal immunization0.2-0.3ml containing peptides70ug/one Mouse. The fourth time in10days:take a Balb/c mice, intraperitoneal immunization0.2-0.3ml, containing peptides100ug/one Mouse. After72hours-96hours,fusion spleens of mice lymphocytes and SP2/0myeloma cells. Use of small peptide-coated ELISA plates selected positive clones repeatedly, and3times by limiting dilution subcloning. Result obtained a specific anti-secretory protein of Marek’s disease virus UL13positive hybridoma cell lines. Named MDV UL13-DE4, MDV UL13-CF8, MDV UL13-FB3, MDV UL13-EC3. This 2monoclonal antibody can specifically react with MDV virus. The development of monoclonal antibodies use for UL13protein function research has laid a solid foundation.2Using monoclonal antibody research MDV UL13To study the expression of the process of viral replication, UL13protein localization in cells. In the Virus replication, UL13protein function and expression contents. Through the use of specific monoclonal antibodies against MDV UL13, Using indirect immunofluorescence assay, Western-blot test, detect CVI988attenuated infection of chicken embryo fibroblasts for the observation of MDV UL13protein detection, to examine the cellular localization of UL13protein. And identified by Western-blot test whether the prokaryotic protein response.A new approach through induced expression of Deng Xufang constructed UL13protein of E. coli. And using mouse antiserum to verify, Makes the purified protein can be used, Opened up new ways of purified protein, in the future provide a new method of purification Protein kinase. And to study its kinase function has laid a good foundation.