不同细胞对柯萨奇病毒A组10型毒株的敏感性及滴度检测影响因素分析  

Analysis of the sensitivity of different cells to coxsackievirus A10 strain and the influencing factors of titer detection

作　　者：高微捷 岳磊[1] 杨婷[1] 宋霞[1] 李华[1] 谢天宏[1] 何鑫 卫星辰 谢忠平[1] GAO Wei-jie;YUE Lei;YANG Ting;SONG Xia;LI Hua;XIE Tian-hong;HE Xin;WEI Xing-chen;XIE Zhong-ping(Key Laboratory for Vaccine Research and Development of Major Infectious Diseases in Yunnan Province,Institute of Medical Biology,Chinese Academy of Medical Sciences,Kunming,Yunnan 650118,China)

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所云南省重大传染病疫苗研发重点实验室,云南昆明650118

出　　处：《中国病毒病杂志》2020年第4期267-271,共5页Chinese Journal of Viral Diseases

基　　金：云南省科技计划项目(2015BC008);中国医学科学院医学与健康科技创新工程项目(2016-I2M-1-019)。

摘　　要：目的探讨不同细胞对柯萨奇病毒A组10型(coxsackievirus A10,CV-A10)毒株的敏感性和滴度检测影响因素,为临床样品分离、病毒培养及保存使用提供基础数据。方法10株已在非洲绿猴肾(African green monkey kidney,Vero)细胞分离并培养2代的CV-A10毒株,检测其在Vero细胞、人胚肺二倍体细胞(human embryonic lung diploid cell,KMB17)、人横纹肌肉瘤(human rhabdomyosarcoma,RD)细胞及人喉癌细胞(human laryngeal carcinoma cell,HeP-2)上的感染性滴度,并对毒株进行蚀斑纯化和冻融处理。结果CV-A10毒株均能适应Vero细胞、KMB17细胞及RD细胞,不能适应HeP-2细胞。RD细胞上第1天即开始病变,第4天达到增值高峰,感染性滴度最高,达到7.75 lgCCID50/ml;KMB17细胞上病变最慢,第3天发生病变,第5天达到增值高峰,滴度为4.85 lgCCID50/ml。毒株在37℃放置3 d感染性滴度明显下降,4℃放置3 d,-30℃和-80℃放置30 d感染性滴度均无影响。结论RD细胞是CV-A10毒株最敏感的细胞,HeP-2细胞不敏感。CV-A10毒株反复冻融次数不宜超过5次,保存温度越低越好并避免4℃和常温下长时间放置。Objective To study the factors influencing the sensitivity and titer detection of different cells to coxsackievirus A10(CV-A10).Methods Ten CV-A10 strains isolated and cultured in African green monkey kidney cells(Vero)for 2 generations were detected on Vero,human embryonic lung diploid cell(KMB17),human rhabdomyosarcoma(RD)and human laryngeal carcinoma cell(HeP-2).Virus strain was subjected to plaque purification and freeze-thaw treatment.Results All CV-A10 strains could adapt to Vero,KMB17 and RD cells,but could not adapt to HeP-2 cells.Viral strains showed the fastest time in RD cells to start lesions on the 1 st day and reached peak on the 4 th day,with the highest infectious titer at 7.75 lgCCID50/ml;the slowest lesions on KMB17 cells that occurred on the 3 rd day and peaked on the 5 th day with titer at 4.85 lgCCID50/ml.The infectious titer of the virus strain was significantly reduced after being placed at 37℃for 3 d,and the infectious titer was not affected by being placed at 4℃for 3 d,and being kept at-30℃and-80℃for 30 d.Conclusions RD cells are the most sensitive cells to CV-A10,and HeP-2 cells are not sensitive.The frequency of repeated freezing and thawing of the virus should not exceed 5 times.The lowest storage temperature of-80℃is better.The virus sample should be avoided for prolonged storage at 4℃and normal temperature.

关 键 词：柯萨奇病毒A组10型 感染性滴度 生长特性 细胞敏感性 VERO细胞 RD细胞 KMB17细胞 

分 类 号：R373.9[医药卫生—病原生物学]