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目的：获得重组ＦＡＳ抗原，用于抗体制备及进一步的功能分析。方法：用ＰＣＲ技术从完整的ＦＡＳｃＤＮＡ克隆上扩增其编码胞外区的部分，将ＰＣＲ产物直接克隆到ｐＧＥＭ－Ｔ载体系统，ＤＮＡ序列分析证实序列完全正确；用ＥｃｏＲⅠ和ＳａｌⅠ将ＦＡＳ胞外区片段切出，定向克隆到经同样酶切处理的ｐＧＥＸ－ＫＧ表达载体，转化大肠杆菌，经优化ＩＰＴＧ的诱导条件，获得了ＦＡＳ胞外区与谷胱甘肽－Ｓ－转移酶的融合蛋白高效表达；用亲合层析法从细菌粗提物中纯化了ＧＳＴ－ＦＡＳ融合蛋白，免疫家兔制备了抗ＦＡＳ抗体。结果：用重组ＦＡＳ为免疫原制备的抗体能诱导Ｕ９３７细胞产生细胞凋亡。结论：重组ＦＡＳ抗原和制备的抗体能用于对ＦＡＳ系统的功能研究
Objective: To obtain recombinant FAS antigen for antibody preparation and further functional analysis. Methods: PCR was used to amplify the part of the extracellular domain from the complete FAS cDNA clone. The PCR product was directly cloned into pGEM-T vector system. DNA sequence analysis confirmed that the sequence was correct. EcoRⅠand SalⅠof FAS extracellular domain The fragment was cut out and cloned into the pGEX-KG expression vector with the same restriction enzyme digestion. The recombinant plasmid was transformed into E. coli. The fusion protein of FAS extracellular domain and glutathione-S-transferase The GST-FAS fusion protein was purified from the crude bacterial extract by affinity chromatography and immunized rabbits to prepare anti-FAS antibody. Results: Antibodies prepared with recombinant FAS as immunogen induced apoptosis in U937 cells. CONCLUSIONS: Recombinant FAS antigens and prepared antibodies can be used for functional studies of the FAS system