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目的制备并鉴定鼠抗博尔纳病病毒核蛋白单克隆抗体。方法重组核蛋白免疫Balb/c小鼠,取其脾细胞与SP2/0骨髓瘤细胞进行常规融合,用间接ELISA法筛选阳性杂交瘤并进行有限稀释法克隆和亚克隆,建立分泌抗核蛋白单克隆抗体的杂交瘤细胞株,纯化单克隆抗体并进行效价测定和特异性鉴定。结果建立了1株能稳定分泌抗核蛋白单克隆抗体的杂交瘤细胞株,命名为2F6E8,重链为IgG2a,轻链为κ。ELISA法测得腹水的效价高达1∶32 000以上。Western-blot和间接免疫荧光实验证实该单克隆抗体能特异性结合重组核蛋白及天然核蛋白。结论成功制备了效价高、特异性好的抗核蛋白单克隆抗体,为建立博尔纳病病毒血清免疫学检测方法奠定了基础。
Objective To prepare and identify murine anti-Burnavirus nucleoprotein monoclonal antibodies. Methods Balb / c mice were immunized with recombinant nucleoprotein. The spleen cells were fused with SP2 / 0 myeloma cells. The positive hybridomas were screened by indirect ELISA and cloned and subcloned by limiting dilution. The hybridoma cell line of the cloned antibody is purified and the titer and specificity of the monoclonal antibody are determined. Results A hybridoma cell line stably secreting monoclonal anti-ribonucleoprotein was established and named as 2F6E8. The heavy chain was IgG2a and the light chain was κ. Ascites titer measured by ELISA up to 1: 32 000 or more. Western-blot and indirect immunofluorescence assay confirmed that the monoclonal antibody can specifically bind to recombinant nucleoprotein and natural nucleoprotein. Conclusion The McAb with high titer and specificity was successfully prepared, which laid the foundation for the establishment of immunological detection of Borna disease virus.