摘要
目的 建立基孔肯雅病毒抗原检测方法.方法 利用重组杆状病毒表达的基孔肯雅病毒全部结构蛋白组成的病毒样颗粒免疫小鼠和兔,制备了基孔肯雅病毒特异性多克隆抗体,建立了检测基孔肯雅病毒抗原双抗体夹心ELISA方法.优化了抗体使用浓度和ELISA反应条件,以灭活基孔肯雅病毒为阳性参考品评价了该方法的检测限和重复性.结果 利用10份模拟阳性血清、90份阴性血清、40份其他病毒感染者急性期血清初步评价该方法的特异性、灵敏性.结果显示特异性为100%,检出限约为10TCID50,板间变异系数小于10%,板内变异系数小于5%.结论 双抗体夹心ELISA方法可以用于检测急性期血清样本中基孔肯雅病毒抗原,为基孔肯雅热的病原学诊断提供了新的手段.
Objective To establish a method for detection of chikungunya virus（CHIKV） antigen.Methods CHIKV virus like particle（VLP）,that contains all structural proteins, was prepared by baculovirus expression system.Mice and rabbits were immunized with the VLP to develop antibodies against CHIKV.A double antibody sandwich ELISA was established for detection of CHIKV antigens.The concentrations of the antibodies used and the reaction conditions were optimized.The detection limit and repeatability of the ELISA was evaluated.Results The sensitivity and specificity was estimated by 10 mimicking CHIKV sera, 90 health person sera,40 other virus infected sera.It was show that the specificity of DAS-ELISA was 100%, the detection limit was 10 TCID50,the coefficients of variation （CV） within plate was 〈5%,the CV of different plates was 〈10%.Conclusions The double antibody sandwich ELISA established in this study can be used to detect the CHIKV antigen in acute phase serum of patient and provide a method for detection of CHIKV.
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
2017年第2期148-152,共5页
Chinese Journal of Experimental and Clinical Virology
基金
国家重点研发计划（2016YFD0500303）
关键词
基孔肯雅病毒
病毒样颗粒
双抗体夹心ELISA
Chikungunya virus
Virus like particle
Double antibody sandwich ELISA
