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抗人纤维蛋白单链抗体 |
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李文平 许 静 隋建华 宋增璇
摘 要 目的：探索抗人纤维蛋白单链抗体-链亲合素融合蛋白的应用前景。方法：构建抗人纤维蛋白单链抗体scFv-8E5基因与链亲和素基因的重组表达载体pSTE2-8E5，表达的融合蛋白可特异性识别人纤维蛋白，同时有生物素的结合位点。转化JM109后用IPTG诱导表达，ELISA观察融合蛋白的稳定性。结果：抗人纤维蛋白单链抗体-链亲合素融合蛋白稳定性好，可耐受至少2 w，4℃保存、数次反复冻融或一定时间的37℃孵育。它能与大鼠和豚鼠的纤维蛋白结合，而与猪纤维蛋白反应不明显。结论：抗人纤维蛋白单链抗体-链亲和素融合蛋白适用于大鼠或豚鼠血栓病模型，是临床前研究的有用材料。
关键词 链亲和素 融合蛋白 抗纤维蛋白
中国图书分类号 R392.2
Stability of anti-human fibrin single-chain antibody: core-streptavidin fusion protein and interaction with other animal fibrin
LI Wen-Ping，XU Jing, SUI Jian-Hua et al.
Institute of Hematology, Chinese Academy of Medical Sciences, Tianjin 300020
Abstract Objective: To explore the clinical use of anti-human fibrin single-chain antibody: core-streptavidin fusion protein in the future. Methods:The author have constructed an anti-human fibrin single-chain antibody: core-streptavidin fusion protein by combining the core-streptavidin sequence with the anti-human fibrin scFv-8E5 gene. The fusion protein expressed by the vector pSTE2-8E5 contained the biotin binding site which could be used for direct detection of fibrin in ELISA with biotinylated horseradish peroxidase. The transformed E. coli JM109 cells were propagated and induced by IPTG. Results:This fusion protein was rather stable; it could tolerate 4℃ storage for at least two weeks, freezing/thawing several times or incubation at 37℃ for a certain time. This fusion protein was also able to bind to rat or guinea pig fibrin, but not to pig fibrin. Conclusion:Anti-human fibrin single-chain antibody: core-streptavidin fusion protein could be taken for developing reliable experimental animal models for the study of thrombus imaging and the targeting of thrombolytic agents.
Key words Streptavidin Fusion protein Anti-fibrin
血栓形成与动脉粥样硬化、栓塞等很多病理生理状态有关,发病率和死亡率均很高。据报道抗人纤维蛋白单链抗体与放射性核素或效应分子结合在血栓性疾病的定位诊断和导向溶栓治疗方面有很好的应用前景［1，2］。Rosebrough等用抗人纤维蛋白与链亲和素交联，产生的融合蛋白在两步法血栓显影中获得良好的效果［3］。本室用基因拼接方法构建了抗人[1] [2] [3] 下一页
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