抗猪囊虫单克隆抗体杂交瘤细胞株的建立和特性鉴定

Establishment of Mouse Hybridoma Cell Lines Secreting Monoclonal Antibody Against Cysticercus Cellulosae Antigen and Characterization

目的 :培养、筛选和克隆鼠源杂交瘤细胞株 ,生产相应特异的针对猪囊虫的单抗并用于临床。方法 :用含有弗氏不完全佐剂和 1单位IFN的猪囊虫囊液皮下注射免疫 7周龄小鼠 3次后 ,无菌取出脾细胞与小鼠骨髓瘤细胞SP2 / 0 ,在含有5 0 %PEG(分子量 4 0 0 0 )的HAT培养液中进行融合 ,在 37℃ ,含 7%CO2 的培养箱中培养 ,随后在正常培养液中进行筛选和克隆化。染色体分析用常规的中期染色体法 ,单抗类型的滴度测定用ELISA法。结果 :用常规的有限稀释法对杂交瘤进行 3次筛选和克隆化后获得 3株杂交瘤细胞株 2H10、5C2和 5H6 ,分别有 97、98和 10 6条染色体 ,均能稳定地分泌各自特异的单抗。 2株单抗为IgG类 ,另 1株为IgM类 ,滴度分别为 1∶10 0 0、1∶80 0和 1∶16 0 0。结论 :用杂交瘤技术 ,我们获得了 3株杂交瘤细胞株 。

Objective:To generate mouse hybridoma cell lines and to induce production of specific monoc1ona1 antibody against cysticercus cellulosae antigen,and eventually being put into practical use clinically Methods:Spleen cells of BALB/C mouse,aged 7wk,immunized subcutaneously 3 times with cysticercus fluid(mixed with Freund's incomplet adjuvant and Interferon)were fused with myeloma cells SP2/O(in the ratio of 5∶1)in the HAT medium containing 50% PEG(MW 4000)and incubated in 37℃,7% CO 2 incubator for several days, and the resulting hybridoma cells were selected and cloned Numerical analysis on Chromosome was performed by using metaphase chromosome technique The type and titre of the McAbs were assayed by ELISA metheds Results:After being selected and cloned 3 times by using routine limited dilution method, 3 hybridama cell lines (2H10,5C2 and 5H6) were generated,which contained 97,98 and 106 chromosomes respectively and secreted steadily specific McAbs Two McAbs belonged to IgG type, and the another one IgM type All the McAbs could specifically bind with the antigen of cysticercus fluid The titres were 1∶1000,1∶800 and 1∶1600 respectively Conclusion:By the hybridoma protocol,we have got 3 hybridoma cell lines that secrete specitic McAbs against cysticercus fluid antigen