流感病毒裂解工艺的优化  

Optimization of procedure for cleavage of influenza virus

作　　者：张赐强[1] 姚为民[1] 刘旭光[1] 孙悦征 赵丹红[1] 曾亮[1] 刘燕[1] 顾建阳[1] 李晓波[1] 郭占龙[1] 杨旭[1] 王亚军[1] 

机构地区：[1]长春生物制品研究所有限责任公司疫苗六室,吉林长春130062

出　　处：《中国生物制品学杂志》2014年第11期1454-1458,1462,共6页Chinese Journal of Biologicals

基　　金：国家科技重大专项(2012ZX09201301-006);吉林省医药产业发展专项资金(YYZX201112)

摘　　要：目的探讨影响流感病毒裂解的因素,优化流感病毒裂解工艺。方法分别在不同裂解时间（Triton X-100裂解1、2、3和4 h）、不同终浓度的裂解剂（终浓度为0.5%、0.6%、0.7%和0.8%的Triton X-100）、不同总蛋白浓度（2 000、3 000和4 000μg/ml）及不同的离子强度[终浓度为0.01、0.05、0.1和0.5 mol/L的PBS（p H 7.2）]下,对流感病毒进行裂解,电镜下观察病毒裂解效果,同时采用免疫单扩散法检测血凝素含量,Lowry法检测总蛋白含量,计算蛋白含量/血凝素含量比值（裂解后病毒纯度）及血凝素回收率,确定最佳裂解工艺参数。结果当裂解前病毒纯化液蛋白含量范围为2 000~3 000μg/ml,PBS终浓度为0.1 mol/L时,利用终浓度为0.7%~0.8%的Triton X-100裂解流感病毒2 h,可获得最佳裂解效果。结论确定了流感病毒最佳裂解参数,优化了流感病毒裂解疫苗生产中的裂解工艺。Objective To investigate the influencing factors and optimize the procedure for cleavage of influenza virus.Methods Influenza virus at various total protein concentrations（2 000,3 000 and 4 000 μg / ml）and ionic strengths[PBS（p H 7. 2）at final concentrations of 0. 01,0. 05,0. 1 and 0. 5 mol / L]was cleaved with Triton X-100 at various final concentrations（0. 5%,0. 6%,0. 7% and 0. 8%）for 1,2,3 and 4 h,observed under electron microscope,then determined for hemagglutinin content by single immunodiffusion test,and for total protein content by Lowry method,based on which the ratio of protein content to hemagglutinin content（purity after cleavage） and recovery rate of hemagglutinin were calculated,and the parameters of cleavage procedure were determined. Results The optimal protein content in purified virus before cleavage was 2 000 ~ 3 000 μg / ml,while the optimal final concentration of PBS was 0. 1 mol / L,the optimal final concentration of Triton X-100 was 0. 7% ~ 0. 8%,and the optimal time for cleavage was 2 h. Conclusion The optimal parameters for cleavage of influenza virus were determined,and the procedure for cleavage of influenza virus was optimized.

关 键 词：流感病毒 TRITON X-100 裂解工艺 优化 

分 类 号：R373.13[医药卫生—病原生物学] R392-33[医药卫生—基础医学]