H5N1亚型禽流感病毒免疫血清精制工艺的建立

目的建立马抗H5N1亚型禽流感病毒免疫血清的精制工艺。方法以硫酸铵盐析法提取免疫马血浆中的IgG抗体,以8～256μg/mgIgG胃蛋白酶(活性单位1:3000)进行消化,以阳离子交换层析柱纯化F(ab’)2抗体,并参照《中国药典》三部(2005版)要求,对试制的样品进行检定。结果经一步50%硫酸铵和多步33%硫酸铵盐析,获得了较为纯净的IgG抗体。8μg胃蛋白酶用量可完全消化1mgIgG抗体分子,阳离子交换方法分离F(ab’)2抗体纯度可达90%以上,高于常规工艺制备的抗体纯度。以此工艺试制的样品,各项质量指标均符合《中国药典》三部(2005版)质量标准。结论已初步建立了马抗H5N1亚型禽流感病毒免疫血清的精制工艺。

Development of A Procedure for Preparation of Purified Horse Immune Serum Against Avian Influenza H5N1 Virus

Objective To develop a procedure for preparation of purified horse immune serum against avian influenza H5N1 virus.Methods The IgG was extracted from horse plasma immunized with avian influenza H5N1 virus by ammonium sulfate precipitation,digested with 8 ～ 256 μg /mg pepsin with an activity of 1:3 000,and purified by cation exchange chromatography.The obtained F(ab')2 antibody was subjected to control tests according to Chinese Pharmacopeia(2005 edition).Results Purified IgG was obtained after one step of precipitation with 50% ammonium sulfate and several steps of precipitation with 33% ammonium sulfate.A portion of 1 mg of IgG was completely digested with 8 μg of pepsin.The purity of F(ab')2 antibody purified by cation exchange chromatography reached more than 90%,which was higher than that by routine procedure.All the quality indexes of immune sera prepared by the developed procedure met the requirements in Chinese Pharmacopeia(2005 edition).Conclusion A procedure for preparation of purified horse immune serum against avian influenza H5N1 virus was preliminarily developed.