Preparation and Functional Characterization of Monoclonal Antibodies against σC Protein of Genotype Ⅳ Avian Reovirus
In order to prepare monoclonal antibodies against σC protein of genotype Ⅳ(GC Ⅳ)avian reovirus(ARV),the recombinant prokaryotic expression plasmid pCold-I-ARV-GC Ⅳ-σC was constructed and transformed into the BL21(DE3)competent cells for protein expression;subsequently,the purified σC protein was immunized to BALB/c mice,and hybridoma cells were prepared by cell fusion,and positive ones were screened by indirect enzyme-linked immunosorbent assay(ELISA);the hybridomas,after identification by indirect immunofluorescence(IFA)and Western blot,was intraperitoneally injected into the mice to prepare monoclonal antibody(mAb)ascites whose titer was measured by ELISA.The results showed that a positive hybridoma cell line(1H4)secreting mAb against ARV GC Ⅳ σC protein was successfully screened;the prepared mAb could react with ARV GC Ⅳ σC protein,and well interact with LMH cells infected with different genotypes of ARV;the mAb heavy chain was identified as IgG1,and the ELISA titer of ascites was 1:256 000.In conclusion,mAb against ARV GC Ⅳ σC protein with good titer and reactivity were successfully prepared in this study,laying a foundation for further researches on the functions of ARV GC Ⅳ σC protein,the pathogenesis of ARV and the development of clinical diagnostic methods.
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