甲型流感病毒在MDCK细胞上培养条件的优化  

Optimization of culture condition for cultivation of influenza virus A on MDCK cell

作　　者：赵晓辉[1] 周大宇[2] 

机构地区：[1]辽宁医学院附属第一医院,锦州121001 [2]辽宁省锦州市疾病预防控制中心

出　　处：《卫生研究》2014年第2期210-212,218,共4页Journal of Hygiene Research

基　　金：国家重大传染病防治科技重大专项(No.2012ZX10004209-001)

摘　　要：目的优化甲型流感病毒在MDCK细胞上的培养条件,提高甲型流感病毒的分离率和监测效果。方法将三种甲型流感病毒液(甲型H1N1、季节性H1N1、季节性H3N2)分别接种于MDCK细胞,通过比较不同的病毒接种量(25、50、75、100、125和150μl/cm2)、不同的TPCK-胰酶添加浓度(1.0、1.5、2.0、2.5、3.0和3.5μg/ml)以及不同的MDCK细胞增毒代次(连续盲传四代),对三种甲型流感病毒易感性的影响,确定其最佳分离效果。结果三种甲型流感病毒:甲型H1N1、季节性H1N1、季节性H3N2的最佳病毒接种量分别为:100、75和75μl/cm2;胰酶最佳使用浓度分别为2.5、2.0和2.0μg/ml;最适宜的细胞增毒传代次数为:一代、二代、二代;低代次的MDCK细胞对甲型H1N1流感病毒的易感性高于季节性H1N1和季节性H3N2;甲型H1N1流感在培养后病毒的HA滴度低于季节性H1N1与季节性H3N2。结论甲型H1N1流感病毒在MDCK细胞上的分离效果低于季节性H1N1与季节性H3N2;季节性H1N1流感病毒和季节性H3N2流感病毒的分离条件基本相同。Abstract：Objective To optimize the cultivation condition of influenza virus A on MDCK cells and to improve the separation rate and the inspected effect of influenza virus. Methods The virus specimens which came from three kinds of the influenza virus A, swine-origin H1N1 ,seasonal H1 N1 and seasonal H3N2, were cultivated by MDCK cells. It will make the most suitable condition by means of comparing the different inoculum size （25,50,75,100,125 and 150 μg/cm2 ） , the different concentration of TPCK-Trypsin （1.0,1.5,2.0,2.5,3.0 and 3.5 μg/ml）as an addition and the different susceptibility of cell generations for serial passage culture of four generations of these three influenza virus. Results Throughout the comparison the three kinds of the influenza virus, swine-origin H1N1 ,seasonal H1N1 and seasonal H3N2, which had the most suitable inoculum size 100,75, 75μl/cm2 and the most suitable concentration of TPCK-Trypsin as an addition2.5,2.0,2.0 p,g/ml and the most suitable generations of cell 1,2,2 respectively. Earlier generations of MDCK cells of the swine-origin H1N1 were more susceptible than the other influenza virus A. The hemagglutination inhibition （HI） of swine-origin H1N1 after cultivated was lower than the other of seasonal H1N1 and seasonal H3N2. Conclusion The separating effect of the swine-origin HIN1 virus in the MDCK cell was inferior to the seasonal H1N1 and seasonal H3N2. The culture condition of seasonal H1N1 and seasonal H3N2 were roughly similar.

关 键 词：甲型流感病毒 MDCK细胞 分离培养 

分 类 号：R511.7[医药卫生—内科学]