创伤弧菌溶细胞素T和B有效联合表位的筛选与鉴定  

Screening and Identification of Efficient T and B Combined Epitopes of Vibrio vulnificus Hemolysin Based on Phage Display Technique

作　　者：王波[1] 丁卉[1] 谢旦立[1] 楼永良[1] 肖美英[1] 庞碧芳[1] 严杰[2] 

机构地区：[1]温州医学院检验医学院浙江省医学遗传学重点实验室,温州325035 [2]浙江大学医学院病原生物学教研室,杭州310031

出　　处：《中国细胞生物学学报》2010年第1期109-114,共6页Chinese Journal of Cell Biology

基　　金：浙江省自然科学资金(No.Y2090468);浙江省科技计划(No.84008012)资助~~

摘　　要：利用生物信息学预测技术成功筛选出含部分创伤弧菌溶细胞素(Vibrio vulnificushemolysin,VvhA)基因片段的T细胞、B细胞联合表位Vvha1、Vvha2、Vvha3和Vvha4,通过噬菌体展示技术成功获得四段表位多肽M13KE-Vvha1、M13KE-Vvha2、M13KE-Vvah3、M13KE-Vvha4,且M13KE-Vvah2、M13KE-Vvha4抗原反应性强于M13KE-Vvha1、M13KE-Vvha3。将四种抗原性多肽经腹腔注射ICR小鼠后,利用流式细胞仪及ELISA方法检测小鼠CD4^+T淋巴细胞亚群及IL-2、IL-4、IL-6、IFN-γ细胞因子时发现,与M13KE-Vvha3相比,经M13KE-Vvha1、M13KE-Vvha2、M13KE-Vvha4作用的小鼠CD4^+T淋巴细胞亚群的变化更为显著。与正常组相比,该三组小鼠的IL-2、IL-4、IL-6、IFN-γ细胞因子分泌量的变化都具有统计学意义;而经M13KE-Vvha3作用的小鼠仅IL-4的改变具有明显统计学意义。因此本研究成功筛选出含部分VvhA片段的T细胞、B细胞联合表位Vvha2、Vvha4,从而为进一步研制多抗原肽(multipleantigenic peptide,MAP)疫苗打下基础。The epitopes of T and B cells in Vibrio vulnificus hemolysin （VvhA） were predicted and analyzed by bioinformatics technique. We discovered four major T and B combined epitopes, named Vvha1, Vvha2, Vvha3 and Vvha4, respectively. The four recombinant epitopes were successfully expressed by phase display system, named M13KE-Vvha1, M13KE-Vvha2, MI3KE-Vvah3, MI3KE-Vvha4, and the antigenicities of M13KE-Vvah2 and M13KE-Vvha4 were better than that of M13KE-Vvha1 and M13KE-Vvha3 through Western blot. The immunogenicities of combined epitopes were further confirmed by flow cytometry and ELISA according to the changes of CD4^＋, CD8^＋ T cells and cytokines, such as IL-2, IL-4, IL-6, IFN-γ. The subgroup of CD4^＋ T cells of ICR mice which were injected intraperitoneally with M13KE-Vvhal, M13KE-Vvha2 and M13KE-Vvha4 changed apparently, and the cytokines also had a significant change compared with cytokines of control groups. But when mice were intraperitoneally injected with M13KE-Vvha3, only IL-4 changed significantly. In conclusion, we had discovered and screened two efficient combined epitopes successfully, Vvha2 and Vvha4, and they could be as candidates of multipleantigenic peptide vaccine.

关 键 词：创伤弧菌 溶细胞素 噬菌体展示 B细胞表位 T细胞表位 

分 类 号：R392[医药卫生—免疫学]