Single-chain antibody streptavidin fusions: tetrameric bifunctional scFv-complexes with biotin binding activity and enhanced affinity to antigen.
Single-chain antibody streptavidin fusions: tetrameric bifunctional scFv-complexes with biotin binding activity and enhanced affinity to antigen.
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单链抗体链霉亲和素融合物：四聚体双功能 scFv 复合物，具有生物素结合活性和增强的抗原亲和力。
DOI：
10.3233/hab-1995-6303
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Stefan Dübel
中科院分区：
文献类型：
--
作者：
Sergey Kipriyanov;Frank Breitling;Melvyn Little;Stefan Dübel
为了增加单链抗体（scFv）对其抗原的亲和力，我们将它们融合到核心链亲和素中。该嵌合蛋白由大肠杆菌载体pSTE（链亲和素标记表达质粒）表达，可形成四聚体复合物，与抗原结合，每个四聚体复合物含有4个生物素结合位点。在羧基末端插入的额外半胱氨酸进一步稳定了该复合物。scFv融合蛋白四聚体可通过亲和层析富集来自质周包涵体的生物素类似物2-亚胺生物素。我们还发现，用生物素化的辣根过氧化物酶染色后，scFv融合蛋白可以在ELISA中直接检测其抗原。由于四聚体结构的亲和效应，抗体复合物的亲和力大大增加。生物素结合位点可用于偶联其他抗体和分子，形成双特异性和双功能试剂。
To increase the avidity of single-chain antibodies (scFv) for their antigen, we have fused them to core-streptavidin. The chimeric protein, expressed by the vector pSTE (plasmid for streptavidin-tagged expression) from Escherichia coli, can form tetrameric complexes, binds its antigen and contains four biotin binding sites per tetrameric complex. An additional cysteine inserted near the carboxy terminus further stabilised the complex. The scFv fusion protein tetramers could be enriched by affinity chromatography using the biotin analog 2-iminobiotin from periplasmic inclusion bodies after refolding. We have also shown that the scFv fusion protein could be used for direct detection of its antigen in ELISA when stained with biotinylated horseradish peroxidase. The affinity of the scFv-antibody complex was substantially increased by avidity effects due to the tetrameric structure. The biotin binding sites may be used for coupling other antibodies and molecules to form bispecific and bifunctional reagents.
影响因子：
56.9
作者：
BIRD, RE;HARDMAN, KD;WHITLOW, M
通讯作者：
WHITLOW, M
DOI：
10.1073/pnas.85.16.5879
发表时间：
1988-08-01
影响因子：
11.1
作者：
HUSTON, JS;LEVINSON, D;OPPERMANN, H
通讯作者：
OPPERMANN, H