EB病毒Rta优势表位抗原的克隆表达及在应用ELISA诊断鼻咽癌中的价值  

Cloning and Expression of Dominant Epitope Antigen of Epstein-Barr Virus Rta Protein and Its Application in the Diagnosis of Nasopharyngeal Carcinoma by ELISA

作　　者：张玲 刘杰[2] 王臣玉[2] 张守信[2] 杨丽萍[2] 危利 王超男 焉丽波 宋少峰 陈蒙蒙 冯晓燕 张贺秋 ZHANG Ling;LIU Jie;WANG Chen-yu;ZHANG Shou-xin;YANG Li-ping;WEI Li;WANG Chao-nan;YAN Li-bo;SONG Shao-feng;CHEN Meng-meng;FENG Xiao-yan;ZHANG He-qiu(Medical Institute of Oriental Ocean(Beijing),Bijing 100071,China;Yantai Yuhuangding Hospital,Shandong Yantai 264000,China;Avioq Biotechnology Co.LTD,Shandong Yantai 264000,China)

机构地区：[1]东方海洋(北京)医学研究院,北京100071 [2]烟台毓璜顶医院,山东烟台264000 [3]艾维可生物科技有限公司,山东烟台264000

出　　处：《现代检验医学杂志》2020年第5期1-4,12,共5页Journal of Modern Laboratory Medicine

基　　金：山东省重大科技创新工程项目(2019JZZY011018)。

摘　　要：目的制备高活性EB病毒立即早期蛋白Rta优势表位抗原,并初步评价该抗原在鼻咽癌(nasopharyngeal carcinoma,NPC)诊断中的价值。方法分析Rta氨基酸序列,选取抗原优势表位Rta(301~440aa),合成目的基因,连接pBVGST-6His载体,构建重组pBV-Rta表达质粒,进行诱导表达获得纯化抗原。采用间接ELISA初步评价优势表位抗原在鼻咽癌诊断中的价值,以纯化的pBV-Rta优势表位抗原为包被抗原,抗人IgA,IgG和IgM为二抗,检测由烟台毓璜顶医院提供的50例经临床病理确诊的NPC患者血清样本和90例健康体检者血清样本。结果获得了高效原核表达的pBV-Rta优势表位抗原(301~440aa),经过小样本验证,确定pBV-Rta具有良好的抗原活性和特异度。放大样本进行检测,基于Rta优势表位抗原的Rta-IgG间接ELISA方法检测灵敏度和特异度分别为73.08%和95.79%。Rta-IgG检测的AUC值为0.860。结论Rta优势表位抗原pBV-Rta是优选的可以用于Rta-IgG检测的抗原,利用该抗原所建立的间接ELISA检测方法可以很好地区分鼻咽癌患者和健康对照。Objective To prepare the dominant epitope antigen of Rta of Epstein Barr virus(EBV)immediate early protein,and evaluate its value in the diagnosis of nasopharyngeal carcinoma(NPC).Methods The amino acid sequence of Rta was analyzed,and the dominant epitope antigen of Rta(301~440aa)was selected to synthesize the target gene.An expression-construct for Rta(301~440aa)was engineered by inserting the corresponding DNA into a pBVGST-6His plasmid.The recombinant pBV-Rta expression plasmid was induced expression to obtain the purified antigen.The value of the dominant epitope antigen in the diagnosis of NPC was preliminarily evaluated by indirect ELISA.The purified pBV-Rta dominant epitope antigen was used as the encapsulated antigen,and the anti-human IgA,IgG and IgM were used as the secondary antibody.The sensitivitiy and specificitiy of pBV-Rta were detected in serum of 50 NPC patients confirmed by clinicopathology and 90 healthy patients tested by Yantai Yuhuangting Hospital.Results The domonant epitope antigen(301~440aa)of pBV-Rta was highly prokaryotic expressedin E.coli.After small sample verification,it was determined that pBV-Rta had good antigenic activity and specificity.The sensitivitiy and specificitiyof Rta-IgG were 73.08%and 95.79%,respectively,by indirect ELISA based on Rta domonant epitope antigen,and the AUC value of Rta-IgG was 0.860.Conclusion The Rta dominant epitope antigen pBV-Rta is the preferred antigen that can be used for Rta-IgG detection.The indirect ELISA detection methods of Rta-IgG established using Rta dominant epitope antigen can be used to distinguish NPC patients from healthy controls.

关 键 词：EB病毒 立即早期蛋白Rta 优势表位抗原 鼻咽癌 

分 类 号：R739.62[医药卫生—肿瘤] R730.43[医药卫生—临床医学]