两种方法检测汉坦病毒滴度的比较研究  

A Comparison of Double-antibody Sandwich ELISA and Direct Immunofluoresence Asssy for Detecting and Titering Hantavirus

作　　者：白露[1] 叶伟[1] 于蒙蒙[1] 张亮[1] 于澜[1] 刘梓谕[1] 吴兴安[1] 徐志凯[1] 张芳琳[1] 

机构地区：[1]第四军医大学微生物学教研室,西安710032

出　　处：《科学技术与工程》2012年第9期2137-2141,共5页Science Technology and Engineering

基　　金：国家自然科学基金(30972590;30970148;30801037);陕西省社发攻关项目(2010k01-174;2010k01-169)资助

摘　　要：比较双抗体夹心ELISA法与直接免疫荧光(DFA)法在检测汉坦病毒灵敏度方面的差别。用汉坦病毒76—118株感染Vero—E6细胞,取不同时间点的病毒感染细胞制备细胞爬片或细胞培养物冻融上清,分别用本室制备的抗汉坦病毒单克隆抗体标记荧光素或辣根过氧化物酶(HRP),建立直接免疫荧光法和双抗体夹心ELISA法检测上述细胞中的病毒抗原,并比较两者的检测灵敏度。用空斑形成实验结果作为金标准,以评价两种方法在检测病毒抗原灵敏度方面的差异。结果:用两种检测方法测定不同时间点的汉坦病毒培养物抗原,双抗体夹心ELISA法不仅检出的时间早,而且其病毒滴度均较IFA法高(P<0.01)。说明ELISA法检测汉坦病毒滴度的灵敏度较IFA法更高,并且操作简单、可重复性好、便于大批量规模化检测,因此更适用于汉坦病毒的检测。To compare the sensitivity between double antibody sandwich ELISA and direct immunofluorescence assay（DFA） for detection of hantavirus,Vero-E6 cells were infected by Hantaan virus strain 76—118.Cell climbing slices or freeze-thaw cell culture supernatant was prepared and fluorescein labeled anti-hantavirus monoclonal antibody or horseradish peroxidase（HRP） labeled monoclonal antibody was used to establish method of double-antibody sandwich ELISA or direct IFA to detect viral antigen in these cells,and plaque reduction nuetralization test was used as control.And then the optimal detection times were identified and the sensitivity of the two methods was compared.It is resulted that virus titer detected by double-antibody sandwich ELISA was obviously higher than that of DFA（P0.01）,and the time point when the virus antigen was detected is more earlier in ELISA than in IFA.It is conclused the sensitivity of double-antibody sandwich ELISA was better than that of direct IFA in detecting hantavirus.And the former method was repeatable and easier to manipulate in large scale.The double-antibody sandwich ELISA is optimal methods for detecting hantavirus titer.

关 键 词：汉滩病毒 病毒滴度 双抗体夹心ELISA 直接免疫荧光 

分 类 号：R373.2[医药卫生—病原生物学]