不同细胞对乙型脑炎病毒蚀斑减少中和试验的影响  

Effect of various cells on plaque reduction neutralization test on Japanese encephalitis virus

作　　者：庞雪[1] 张嵘[1] 姚宇[1,2] 孙非非[1,2] 李鹤[2,3] 

机构地区：[1]沈阳药科大学生命科学与生物制药学院生物工程教研室,辽宁沈阳110016 [2]辽宁成大生物股份有限公司质检部,辽宁沈阳110176 [3]辽宁大学生命科学学院,辽宁沈阳110036

出　　处：《中国生物制品学杂志》2016年第2期140-143,共4页Chinese Journal of Biologicals

摘　　要：目的分别使用不同细胞测定乙型脑炎病毒(Japanese encephalitis virus,JEV)滴度及乙型脑炎灭活疫苗效价,分析不同细胞对检测结果的影响。方法将JEV P3株接种至长满单层的MDBK、Vero、LLC-MK2、BHK-21细胞的6孔板,采用蚀斑试验测定JEV的PFU;将乙型脑炎灭活疫苗(Vero细胞)免疫小鼠,取静脉血,分离血清,经56℃灭能30 min后,与JEV P3株混合,接种至长满单层的Vero、BHK-21、LLC-MK2细胞的6孔板,采用蚀斑减少中和试验检测乙型脑炎灭活疫苗的效价。结果 JEV在4种细胞上均可产生细胞病变(cytopathic effect,CPE),但在MDBK细胞上不形成蚀斑,病毒感染的Vero、LLC-MK2、BHK-21细胞的收获液病毒滴度分别为10.02、9.42和9.21 lg PFU/ml;3种细胞测定乙型脑炎灭活疫苗效价T值Vero>LLC-MK2>BHK-21,其中Vero细胞与BHK-21和LLC-MK2细胞比较,蚀斑减少率及对应T值差异有统计学意义(P<0.05),LLC-MK2与BHK-21细胞比较,蚀斑减少率及对应T值差异无统计学意义(P>0.05)。结论 4种细胞感染JEV后均可产生CPE,但MDBK细胞不能形成蚀斑;不同细胞测定疫苗效价,蚀斑减少率及对应T值也不同,其中Vero细胞测定T值最大,BHK-21细胞测定T值最小。Objective To determine plaque forming unit （PFU） of Japanese encephalitis virus （JEV） and the titer of inactivated JE vaccine by using various cells and analyze the effect of the cells on determination result of vaccine titer. Methods JEV P3 strain was inoculated onto the monolayer of MDBK, Vero, LLC-MK2 and BHK-21 cells, and determined for PFU by plaque assay. Mice were immunized with inactivated JE vaccine （Vero cells）, of which the serum samples were collected, inactivated at 56 ~C for 30 rain, mixed with JEV P3 strain, inoculated onto monolayer of Vero, BHK-21 and LLC-MK2 cells respectively, and derermined for vaccine titer by plaque reduction neutralization test. Results JEV eaused cytopathic effect （CPE） of all the four kinds of cells, while formed no plaque on MDBK cells. The virus titers in harvests of Veto, LLC-MK2 and BHK-21 cells were 10. 02, 9. 42 and 9. 21 PFU/ml respectively. In the turn of T values of titers of inactivated JE vaccine, the eells were Vero, LLC-MK2 and BHK-21 cells. The plaque reduction rate and T value showed significant difference in Vero and LLC-MK2 cells （P 〈 0. 05）, while showed no significant difference in LLC-MK2 and BHK-21 cells （P 〉 0. 05）. Conclusion JEV caused CPE in all the cells, while formed no plaque on MDBK cells. The plaque reduction rates and the corresponding T values of various cells for determination of vaccine titer were different, of which the T value was the highest in Vero cells, and the lowest in BHK-21 cells.

关 键 词：乙型脑炎病毒 蚀斑试验 BHK-21细胞 VERO细胞 LLC-MK2细胞 MDBK细胞 

分 类 号：R373.31[医药卫生—病原生物学] R392-33[医药卫生—基础医学]