幽门螺杆菌尿素酶B单克隆抗体的制备与鉴定  

Preparation and Identification of Monoclonal Antibody Against Urease B Subunit from Helicobacter pylori

作　　者：陶好霞[1] 刘向昕[1] 张兆山[1] 展德文[1] 袁盛凌[1] 刘纯杰[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071

出　　处：《生物技术通讯》2007年第2期246-248,共3页Letters in Biotechnology

基　　金：国家高技术研究发展计划项目(2001AA215161;2004AA215213)

摘　　要：目的:制备稳定分泌抗幽门螺杆菌尿素酶B单克隆抗体(mAb)的杂交瘤细胞系,并对其分泌的mAb进行鉴定。方法:用初步纯化的重组幽门螺杆菌尿素酶B免疫BALB/c小鼠,利用杂交瘤技术制备抗尿素酶B的mAb,用间接ELISA检测mAb的特异性和亲和力,检测mAb腹水效价,鉴定Ig亚类并测定其抗原决定簇。结果:获得8株能稳定分泌抗尿素酶B的mAb杂交瘤细胞系,这8株单抗与能产生尿素酶的小肠结肠耶尔森氏菌、肺炎克雷伯氏菌和普通变形杆菌均无交叉反应,相对亲和力为1.13×10-8～4.66×10-10,腹水mAb效价可达2×104～3.2×105。其中2株单抗属IgG1亚类,3株单抗属IgG2a亚类。8株单抗分属于3种不同的抗原决定簇。结论:获得了IgG1和IgG2a类型的针对3种不同抗原决定簇的特异性幽门螺杆菌尿素酶B的mAb,为进一步用于幽门螺杆菌的临床诊断和实验研究创造了条件。Objective： To establish the specific hybridoma cell lines that secreted stably monoclonal antibody（mAb） against urease B subunit（UreB） from Helicobacter pylori. Methods： Female BALB/c mice were immunized with purified recombinant UreB from H.pylori and mAb against UreB was prepared with hybridoma cell fusion technique. Indirect enzyme-linked immunosorbent assay（ELISA） was used to evaluate the affinity and titers of mAbs, determine their specificity, immunoglobulin subtype and identify their antigen determinants. Results： Eight stably secreting anti-UreB mAb cell lines were acquired. Cross-reactivity with Yersinia enterocolitica, KlebsieUa pneumoniae and Proteus vulgaris, which also produce urease, was not detected. Their relative affinities were between 1.13×10^-8-4.66×10^-10. The titers of mAb in ascites were from 2×10^4-3.2×10^5. Two mAb cell lines secreted IgG1 subtype antibodies and three cell lines IgG2a antibodies. All eight antibodies belonged to three different antigen determinants. Conclusion： The specific anti-UreB mAbs of IgG1 and IgG2a subtypes were produced, which will be significant for clinical diagnosis and fundamental research in H.pylori .

关 键 词：尿素酶B 单克隆抗体 幽门螺杆菌 

分 类 号：Q813.2[生物学—生物工程]