| 单链抗体-碱性磷酸酶融合蛋白的制备 |
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| 引用本文： | 许静,刘俊霞,李侠,宋增璇. 单链抗体-碱性磷酸酶融合蛋白的制备[J]. 中国免疫学杂志, 2003, 19(10): 704-706 |
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| 作者姓名： | 许静 刘俊霞 李侠 宋增璇 |
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| 作者单位： | 中国医学科学院血液学研究所实验血液学国家重点实验室,天津,300020 |
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| 基金项目： | 国家自然科学基金资助 (批准号 3 0 170 3 49) |
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| 摘 要： | 目的：制备单链抗体-碱性磷酸酶融合蛋白。方法：把碱性磷酸酶编码区插入pSTE2-8E5质粒DNA，用scFvC66的重链和轻链可变区DNA取代scFv-8E5的重链和轻链可变区DNA，构成表达载体pSTE2-C66-Ap在大肠杆菌中表达，用聚丙烯酰胺凝胶电泳和免疫印迹法分析产物活性。结果：获得一个分子量为75kD的scFvC66-Ap融合蛋白，可结合来自KGla细胞裂解物中的一个分子量约60kD的蛋白质带，其结合功能可通过其碱性磷酸酶活性直接测定。结论：建立了一个用大肠杆菌制备单链抗体-碱性磷酸酶融合蛋白的方法，它可能取代常规的碱性磷酸酶标记方法而用于免疫检测。
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| 关 键 词： | 单链抗体 碱性磷酸酶 融合蛋白 |
| 文章编号： | 1000-484X(2003)10-0704-03 |
Production of single chain antibody-alkaline phosphatase fusion protein |
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| XU Jing,LIU Jun-Xia,LI Xia et al.Chinese Academy of Medical Sciences. Production of single chain antibody-alkaline phosphatase fusion protein[J]. Chinese Journal of Immunology, 2003, 19(10): 704-706 |
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| Authors: | XU Jing LIU Jun-Xia LI Xia et al.Chinese Academy of Medical Sciences |
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| Affiliation: | XU Jing,LIU Jun-Xia,LI Xia et al.Chinese Academy of Medical Sciences,Institute of Hematology,State Key Laboratory of Experimental Hematology,Tianjin 300020,China |
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| Abstract: | Objective:Making single chain antibody(scFv)-alkaline phosphatase(Ap)fusion protein.Methods:An expression vector pSTE2-C66-Ap was constructed by sequentially inserting the Ap coding region into plasmid pSTE2-8E5 and replacing the VH-VL fragment of 8E5 by VH-VL fragment of C66.The fusion protein scFvC66-Ap was expressed in E.coli.and analysed by SDS-PAGE and immunoblotting.Results:Have obtained the scFvC66-Ap fusion protein with a molecular weight of 75 kD.It bound a 60 kD molecule from KG1a cell proteins on immunoblotting membrane detected directly by Ap enzymatic activity.Conclusion:A method permits the production of scFv-Ap conjugates in E.coli.which can replace conventionally prepared Ap-labeled antibodies in immunoassays. |
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| Keywords: | Single chain antibody Alkaline phosphatase Fusion protein |
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