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目的制备马抗H7N9亚型流感病毒精制免疫球蛋白,并评价其对BALB/c小鼠的治疗效果。方法应用昆虫细胞-杆状病毒表达系统分别构建的表达H7N9亚型流感病毒HA、NA及M1蛋白的病毒样颗粒制备抗原,免疫健康马匹,当抗体效价达1∶10 000以上时,采血,对血清进行粗处理;采用亲和层析的方法获得纯化的马抗H7N9免疫球蛋白F(ab’)_2。对纯化的免疫球蛋白进行蛋白含量测定和质量检测。用H7N9亚型流感病毒BALB/c小鼠感染模型评价F(ab’)_2的治疗效果。结果免疫马匹血清中产生高滴度的中和抗体,获得的纯化马抗H7N9免疫球蛋白F(ab’)_2中和抗体效价达1∶5 120,质量检测合格。攻毒4 h后给药0.4和0.2 mg,小鼠存活率均达100%。结论获得了安全、高效的马抗H7N9亚型流感病毒精制免疫球蛋白F(ab’)_2,为人禽流感治疗用制剂的研发提供了参考。
Objective To prepare equine anti-H7N9 subtype influenza virus purified immunoglobulin and evaluate its therapeutic effect on BALB / c mice. Methods The virus-like particles expressing HA, NA and M1 protein of H7N9 subtype influenza virus constructed respectively by the insect cell-baculovirus expression system were used to prepare antigens and immune healthy horses. When the antibody titer was more than 1:10 000, The serum was subjected to rough treatment. The purified horse anti-H7N9 immunoglobulin F (ab ’) 2 was obtained by affinity chromatography. The purified immunoglobulin protein content and quality testing. The therapeutic effect of F (ab ’) 2 was evaluated using a H7N9 subtype influenza virus BALB / c mouse infection model. Results High titer of neutralizing antibody was produced in the serum of immunized horses. The titer of purified neutralizing antibody of horse anti-H7N9 immunoglobulin F (ab ’) _ 2 reached 1:5 120, and the quality test was qualified. Four and four hours after challenged with 0.4 and 0.2 mg, the survival rate of mice reached 100%. Conclusions A safe and efficient anti-H7N9 subtype influenza virus purified immunoglobulin F (ab ’) _ 2 was obtained, which provided a reference for the development of human bird flu therapeutic formulations.