【摘要】 目的 :制备抗禽流感病毒 (AIV)H9亚型血凝素蛋白的单克隆抗体 (mAb)。方法 :以AIVH9亚型油乳剂灭活疫苗作为免疫原 ,免疫 8wk龄BALB/c小鼠。采用淋巴细胞杂交瘤技术制备抗AIVH9亚型血凝素蛋白的mAb ;采用ELISA和血凝抑制试验(HI)检测腹水mAb的效价 ;采用ELISA、HI、免疫荧光染色 (IF)及Westernblot鉴定mAb的特异性。结果 :获得 3株可稳定分泌特异性mAb的杂交瘤细胞株 2A3、2H1和 1C8,其腹水mAb的ELISA效价依次为 1× 10 7、1× 10 5和 5× 10 6,血凝抑制效价为 1× 2 8～ 1× 2 13 ;3株mAb的Ig亚类均为IgG1。以mAb 2H1进行Westernblot的结果显示 ,该mAb能与AIV的Mr 为 75 0 0 0的蛋白条带起反应 ,表明其是针对AIVH9亚型血凝素蛋白的mAb。与 32株AIVH9亚型国内分离株进行血凝抑制试验表明 ,mAb 2H1具有良好的广谱性。结论 :成功地制备了抗AIVH9亚型血凝素蛋白的mAb ,为AIV的抗原性分析、血清学诊断、疫苗质量的监测及流行病学调查等奠定了基础更多还原

【Abstract】 AIM: To prepare monoclonal antibodies （mAb） against the hemagglutinin（HA） of H9 subtype of avian influenza virus （AIV）. METHODS: 8 week-old female BALB/c mice were immunized with the inactivated vaccine of H9 subtype of AIV. Splenocytes from the immunized mice were fused with Sp2/0 myeloma cells, and positive hybridoma clones were screened by indirect ELISA and hemagglutination inhibition test （HI）. The specificity of the mAb was characterized by ELISA, HI test, indirect immunofluorescence （IF） staining and Western blot. RESULTS: Three hybridoma cell lines named 2H1, 2A3 and 1C8 against HA of AIV H9 were obtained. The HI titers of 3 mAbs were 1×2⁸-1×2¹³, and the ELISA titers were 1×10⁷, 1×10⁵ and 5×10⁶, respectively. The immunoglobulin subclasses of all 3 mAbs were IgG1. Western blot analysis confirmed that mAb 2H1 could recognize HA and reacted to 31 out of 32 isolates of H9 subtype of AIV. CONCLUSION: Three mAbs recognizing HA of H9 subtype of AIV were obtained, which may provide an useful tool for the antigenic analysis, the serological diagnosis, the epidemiological survey and the evaluation of AIV vaccine.更多还原