摘要
目的应用复合凝胶Capto Core700作为纯化介质去除狂犬病疫苗中的Vero细胞宿主蛋白和宿主细胞DNA等杂质。方法将Vero细胞接种至微载体,在celligen plus生物反应器中培养,按MOI=0.002接种狂犬病病毒,连续制备10批狂犬病疫苗原液。经过滤及浓缩后加入β-丙内酯,于2~8℃灭活24 h,获得病毒灭活液。上样至装载复合凝胶Capto Core700的纯化柱进行层析纯化,并按《中国药典》三部(2015版)的方法检测宿主细胞蛋白去除率、宿主细胞DNA去除率、狂犬病病毒糖蛋白回收率及总蛋白去除率。同时与传统Sepharose系列凝胶纯化方法进行比较。结果 10批疫苗纯化液的宿主细胞蛋白去除率为58.3%~63.2%,宿主细胞DNA去除率均>90%,糖蛋白回收率范围为52.3%~74.5%,蛋白去除率范围为7.22%~11.76%。与传统Sepharose系列凝胶纯化方法比较,差异较小,且Capto Core700的上样量为3倍柱体积,高于Sepharose凝胶(15%)。结论 Capto Core700凝胶可有效去除狂犬病疫苗中的杂质,缩短了纯化工艺时间。
Objective To remove the foreign matters such as residual Vero cell host protein and host cell DNA from rabies vaccine by using Capto Core700. Methods Vero cells were inoculated onto microcarriers and incubated in celligen plus bioreactor, to which rabies virus was inoculated at MOI of 0. 002. Ten consecutive batches of bulk of rabies vaccine were prepared, filtrated, concentrated, and inactivated with β-propiolactone at 2-8 ℃ for 24 h, then purified with complex media Capto Core700. The purified virus liquid was determined for removal rates of host cell protein and host cell DNA,recovery rate of rabies virus glycoprotein and protein removal rate according to the requirements in Chinese Pharmacopeia（VolumeⅢ, 2015 edition）. Results The removal rates of host cell protein in ten batches of purified vaccine were 58. 3%-63. 2%, while those of host cell DNA were more than 90%, the recovery rates of glycoprotein were 52. 3%-74. 5%, and the protein removal rates were 7. 22%-11. 76%, which showed no significant difference with those of vaccine purified by traditional Sepharose gel chromatography. However, the sample load for purification with Capto Core700 was 3 times of column volume, which was larger than that by Sepharose gel chromatography（15%）. Conclusion The foreign matters in rabies vaccine was effectively removed by using Capto Core700 gel, while the time for purification was shortened.
作者
李旭
徐威
于海
陈志成
LI Xu;XU Wei;YU Hal;CHEN Zhi-cheng(Liaoning Chengda Biological Co., Ltd., Shengyang 100179, Liaoning Province, China)
出处
《中国生物制品学杂志》
CAS
CSCD
2018年第5期538-542,共5页
Chinese Journal of Biologicals
