抗猪囊尾蚴KD26抗原单克隆抗体杂交瘤细胞株建立和特性鉴定

目的: 建立针对猪囊尾蚴KD26抗原的单克隆杂交瘤细胞株,生产相应的单克隆抗体,用于囊虫病免疫保护机制的基础研究和囊虫病临床免疫诊断的应用。方法: 用SDS-PAG E电泳,分离纯化制备猪囊尾蚴KD26抗原免疫6周龄BALB/C小鼠3次,无菌取出脾细胞与小鼠骨髓瘤细胞SP2/O(比率10:1),在50% PEG (4 000MW)作用下进行细胞融合,随后在HT培养液中进行选择性培养、筛选和克隆;常规的中期染色体法分析染色体,单克隆抗体鉴定采用免疫双扩散法测定其免疫球蛋白亚类,采用ELISA法测定单抗效价并作特异性鉴定。结果: 用常规的有限稀释法对杂交瘤细胞进行了3次筛选和克隆化后,获得了1株杂交瘤XH,染色体数为96,能稳定分泌抗猪囊尾蚴KD26单抗,单抗类型为IgG1,抗体滴度为1:1 000,其与血吸虫、弓形虫、细粒棘球绦虫抗原均不发生交叉反应。结论: 应用杂交瘤技术获得了1株能稳定分泌高滴度抗猪囊尾蚴KD26抗原的单克隆抗体细胞株。

Establishment and characterization of mice hybridoma cell line secreting monoclonal antibody against cysticercus cellulosae KD26 antigen

Objective: To develop and identify the monoclonal antibody(McAb) against cysticercus cellulosae KD26 antigen,so as to provide useful help and further study.Methods: BALB/C mice were immunized 3 times with KD26 antigen of cysticercus cellulosae by SDS-PAGE(mixed with Freund's incomplet adjuvant),and the spleen cells of immunized mice were fused with myeloma cell line SP2/0(in the ratio of 10:1) in the HAT medium containing 50% PEG(4 000 MW),then incubated,selected and cloned.Numerical analysis on chromosome was performed by using metephase technique.The type and titer of the McAb was assayed by agar double diffusion technique and ELISA methods.Results: After being selected and cloned 3 times by using routine limited dilution method,one hybridoma cell line XH was generated,which contained 96 chromosomes and could steadily secrete specific McAb.The McAb belonged to IgG1 type.The titer was 1:1 000.The McAb had no cross reactions with the antigens of Schistosome japonicum,Toxoplasma gondii and Echinococcus granulosus.Conclusions: By the hybridoma technique,we have established the hybridoma cell line which can secrete specific and high titers McAb against cysticercus cellulosae KD26 antigen.