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目的探讨结核分枝杆菌热休克蛋白(TBhsp)和结核分枝杆菌T细胞刺激表位(MT)在肝细胞再生磷酸酶-3(PRL-3)单克隆抗体(mAb)制备过程中的佐剂作用。方法构建对照质粒pET28a-PRL-3和佐剂-PRL-3融合蛋白表达质粒pET28a-PRL-3-MT、pET28a-TBhsp-PRL-3和pET28a-TBhsp-PRL-3-MT,并纯化其表达蛋白,分别免疫BALB/c小鼠,ELISA检测并比较各组的抗血清效价,选取效价最高的小鼠,采用杂交瘤技术制备PRL-3 mAb,并进行类和亚类鉴定。结果成功构建了上述4种PRL-3相关的重组表达质粒,并表达纯化出相应的融合蛋白,其中PRL-3-MT融合蛋白免疫组抗体效价高于其他组;用该组小鼠进行细胞融合并筛选获得均为IgG类的10株分泌PRL-3 mAb的细胞株。结论 MT在PRL-3 mAb制备中可以发挥较为明显的免疫佐剂作用。
Objective To explore the adjuvant effect of TBhsp and MT Mycobacterium tuberculosis T cell stimulating epitope (MT) in the preparation of monoclonal antibody (mAb) of hepatocyte-regenerating phosphatase-3 (PRL-3) effect. Methods The expression plasmids pET28a-PRL-3-MT, pET28a-TBhsp-PRL-3 and pET28a-TBhsp-PRL-3-MT of the control plasmid pET28a-PRL-3 and adjuvant-PRL- The BALB / c mice were immunized respectively. The antiserum titers of each group were detected by ELISA. The mice with the highest titer were selected and PRL-3 mAbs were prepared by hybridoma technique. Results The four PRL-3 recombinant plasmids were successfully constructed and the corresponding fusion proteins were purified. The antibody titer of the PRL-3-MT fusion protein immunized group was higher than that of other groups. 10 strains of secreting PRL-3 mAbs that are both IgG class were obtained by fusion and screening. Conclusion MT can play a more significant immune adjuvant in the preparation of PRL-3 mAb.