癌胚抗原单链抗体—链亲和素融合蛋白的功能鉴定

目的：用ｐＥＴ２１原核表达系统表达癌胚抗原单链抗体－链亲和素融合蛋白。方法：融合表达载体转化大肠杆菌菌株ＨＭＳ１７４（ＤＥ３），用１ｍｍｏｌ／Ｌ的ＩＰＴＧ诱导表达，表达产物行亲和印迹及免疫斑点分析。结果：ＳＤＳ－ＰＡＧＥ表明，融合蛋白分子质量约５７ｋｕ，亲和印迹及免疫斑点证实融合蛋白具有结合ＣＥＡ与生物素的双特异性。结论：原核表达系统可以表达具有双特异性的融合蛋白，该融合蛋白利用生物素标记的辣根过

Functional identification of the bispecific fusion protein of CEA singlechain antibody and streptavidin

AIM: To express a fusion protein of carcinoembryonic antigen (CEA) singlechain antibody fragment of variable region(ScFv) and streptavidin by pET21 prokaryotic expression system. METHODS: The E.coli.strain HMS174(DE3) was transformed by the fusion expression vector pACS,then induced to express target protein by 1 mmol/L IPTG.The extract from the transformed strain was analyzed by protein affinity blot and immunological dot assay. RESULTS: SDSPAGE demonstrated that the apparent molecular weight of the target protein was about 57 ku, and affinity blot and immunological dot proved that the fusion protein had bispecific activity and could bind CEA and streptavidin respectively. CONCLUSION: The bispecific fusion protein could be expressed by prokaryotic expression system and used to test CEA directly by biotinated horseradish peroxidase(HRP).