RTCA实时定量动态监测HCMV在MRC-5细胞上的增殖的研究  

Monitoring of HCMV replication with real-time cell assay on MRC-5

作　　者：张业婷 钟峰[1] 姚瑶[1] 赵俊[1] 俞海洋[1] 张文昌[1] 陈敬贤[1] 王明丽[1] 

机构地区：[1]安徽医科大学微生物学教研室,合肥230032

出　　处：《安徽医科大学学报》2016年第7期935-939,共5页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:30872253);安徽高校省级自然科学研究项目(编号:KJ2012ZD08)

摘　　要：目的采用实时细胞分析(RTCA)系统监测人巨细胞病毒(HCMV)在MRC-5细胞株上的复制增殖过程,评价此方法在动态监测病毒增殖中的应用价值。方法应用RTCA动态观察MRC-5细胞在不同阶段的生长指数(CI),选择合适的细胞浓度进行HCMV增殖的研究,同时采用传统的空斑形成试验和间接免疫荧光实验作为该方法学的比较和结果验证。结果选择MRC-5最适细胞浓度为0.5×105个/孔;与空斑试验及免疫荧光结果比较,半数细胞指数CI值对应时间(CIT50)与病毒初始滴度有良好的线性关系;同时CI值与病毒滴度随着时间的持续,显示出明显的负相关性(r=-0.977,P<0.05)。结论 RTCA技术可实时监测记录细胞生长状态,反应细胞贴壁、伸展,同时能实时定量监测HCMV增殖复制状况,数据客观可靠,为今后基础实验研究提供了新的实验技术和方法。Objective To detect the human cytomegalovirus（ HCMV） replication in MRC-5 cells by applying realtime cell assay（ RTCA） system,and evaluate the applicable value of this method in the dynamic monitoring of viral replication. Methods RTCA was applicated to monitor cell index（ CI） in different growth stages. The optimal concentration of cell was selected to study HCMV replication. Additionally,traditional plaque formation assay and immunofluorescence assay were used to compare and validate the result of RTCA system. Results The most suitable cell concentration was 0. 5 × 105 cells per plate. Compared with the plaque formation assay and immunofluorescence assay,the value of half cell index（ CIT50） had a satisfactory linear correlation with the initial virus titer. The RTCA system showed a significant negative correlation with time continuation for the detection of cell index and the virus titer（ r =- 0. 977,P〈 0. 05）. Conclusion RTCA technique is successfully established to monitor the growth status of the cells in real time,which might reflect the cell＇s adhesion and spreading. The RTCA system could timely monitor HCMV titer with cell index. All the data are solid and reliable. The method would be helpful for future basic experiments on creating new experimental techniques and methods.

关 键 词：实时细胞检测技术 人巨细胞病毒 增殖 

分 类 号：R373.9[医药卫生—病原生物学]