论文部分内容阅读
目的制备抗肿瘤坏死因子相关凋亡诱导配体细胞外段(sTRAIL)的单克隆抗体。方法用重组sTRAIL免疫BALB/c小鼠,取脾细胞与SP2/0细胞融合,捕获ELISA筛选阳性克隆,制备腹水抗体。间接ELISA测定单抗效价,并进行单抗亚类、特异性鉴定及杂交瘤细胞染色体和单抗识别位点分析。将杂交瘤细胞体外连续培养3个月及冻存6个月后,检测细胞培养上清的效价。结果3株杂交瘤细胞分泌的单抗均为IgG1型,腹水单抗效价均高于10-6,杂交瘤细胞染色体数介于94～98条之间,均识别sTRAIL分子上线性表位。细胞体外连续培养3个月及冻存6个月后的上清效价保持稳定。结论已成功制备了3株可稳定分泌抗sTRAIL单克隆抗体的杂交瘤细胞,为TRAIL的定性定量检测及组织表达分析奠定了基础。
OBJECTIVE: To prepare monoclonal antibodies against the extracellular domain of apoptosis inducing ligand (sTRAIL) associated with tumor necrosis factor. Methods BALB / c mice were immunized with recombinant sTRAIL. The spleen cells were fused with SP2 / 0 cells and the positive clones were screened by ELISA to prepare ascitic fluid antibodies. Indirect ELISA was used to measure the titer of McAb. The McAbs were identified by their specificity, specificity and chromosomal and monoclonal antibody recognition sites. The hybridoma cells were cultured in vitro for 3 months and frozen for 6 months, and the titer of the cell culture supernatant was detected. Results The monoclonal antibodies secreted by all the three hybridoma cells were of IgG1 type. The titer of ascites was higher than 10-6. The number of chromosomes of hybridomas was between 94 and 98, both of which were linear epitopes on sTRAIL. The cells were cultured continuously for 3 months in vitro and frozen for 6 months after the supernatant titer remained stable. Conclusion Three hybridoma cells that can stably secrete anti-sTRAIL monoclonal antibody have been successfully prepared, which lays the foundation for the qualitative and quantitative detection of TRAIL and the tissue expression analysis.