BIFUNCTIONAL AND MULTIMERIC COMPLEXES OF STREPTAVIDIN FUSED TO SINGLE-CHAIN ANTIBODIES (SCFV)
BIFUNCTIONAL AND MULTIMERIC COMPLEXES OF STREPTAVIDIN FUSED TO SINGLE-CHAIN ANTIBODIES (SCFV)
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DOI：
10.1016/0022-1759(94)00257-w
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发表时间：
1995-01-27
影响因子：
2.2
通讯作者：
LITTLE, M
中科院分区：
文献类型：
--
作者：
DUBEL, S;BREITLING, F;LITTLE, M
具有确定化学计量的多价和多特异性抗体可以为生物学和医学研究以及癌症的诊断和治疗提供有价值的工具。因此，我们将单链抗体（scFv）与核心链霉抗生物素蛋白融合。由载体pSTE-215（用于链霉亲和素标记表达的质粒）表达的这种嵌合蛋白可以形成四聚体复合物，结合抗原并含有可用于进一步复合物形成的生物素结合位点。在羧基末端附近插入额外的半胱氨酸以促进共价连接的双功能分子的构建。用生物素类似物进行亲和层析，可得到纯化的scFv融合蛋白。我们还表明，scFv融合蛋白可以用于直接检测其抗原在ELISA和Western印迹时，用生物素化辣根过氧化物酶染色。
Multivalent and multispecific antibodies with defined stoichiometry could provide valuable tools for biological and medical research and for the diagnosis and therapy of cancer. We have therefore fused single chain antibodies (scFv) with core-streptavidin. This chimeric protein, expressed by the vector pSTE-215 (plasmid for streptavidin tagged expression), can form tetrameric complexes, binds antigen and contains the biotin binding site which may be used for further complex formation. An additional cysteine was inserted near the carboxy terminus to facilitate the construction of covalently linked bifunctional molecules. The scFv fusion protein could be purified by affinity chromatography using biotin analogues. We have also shown that the scFv fusion protein could be used for direct detection of its antigen in ELISA and Western blots when stained with biotinylated horseradish peroxidase.