抗传染性法氏囊病毒VP2蛋白单克隆抗体的制备及鉴定OA北大核心CSTPCD

Infectious bursal disease(IBD)is a highly infectious disease caused by Infectious bursal disease virus(IBDV)and the characteristic lesions included atrophy of bursa of Fabricius and bleeding of leg muscles.At present,the rapid spread of a novel IBDV variant has caused great economic losses to poultry industry in China.Therefore,it is significant to develop an effective immunological detection method for rapid diagnosis of IBD.The RT-PCR product of VP2 gene segment was amplified from the RNA extract of the novel variant IBDV-LY21/2 strain.The recombinant prokaryotic expression plasmid pCold-Ⅰ-IBDV-VP2 was transformed into BL21(DE3)cells for protein expression.The expressed recombinant protein was examined by Coomassie blue staining and Western blot.The VP2 protein was expressed in large quantities and purified for preparation of monoclonal antibodies(MAbs).The molecular weight of the recombinant protein His-VP2 was about 50 kDa and its concentration was 8 mg/mL.Then the purified His-VP2 was used to immunize BALB/c mice and the murine serum samples were titrated in indirect ELISA.As a result,Three hybridoma cell lines(1G10F12,3E3E9 and 4D12G12)were obtained after two cycles of subcloning.The heavy chains of these three MAbs were IgG1 and their light chains were κ.The MAbs 1G10F12 and 3E3E9 reacted with the recombinant protein Myc-VP2 in Western blot and IFA while the 4D12G12 showed reactivity in IFA.This study laid a foundation for development of diagnostic methods for IBD and provided a good tool for further study of the function of VP2 protein.