【摘要】 目的将能与血小板膜糖蛋白 a特异结合的单克隆抗体 SZ- 2 1构建成单链抗体 ,为其临床应用奠定基础。方法通过逆转录及多聚酶链反应 ,扩增并克隆 SZ- 2 1的可变区基因 VH、VL ,经测序后构建 SZ- 2 1单链抗体 (SZ- 2 1Sc Fv) ,在大肠杆菌中进行表达。EL ISA和 Western印迹检测其与血小板的结合特性。结果 VH、VL 可变区基因符合小鼠抗体可变区特征 ,SZ- 2 1Sc Fv基因拼接正确。表达产物除少量为分泌型外 ,主要以包涵体形式存在 ,表达量占菌体蛋白的 2 1%。经过变性和复性后 ,该单链抗体保留了与血小板特异结合的能力。结论成功表达了 SZ- 2 1单链抗体 ,该小分子抗体具有特异结合血小板的能力 ,有望用于抗血栓治疗。更多还原

【Abstract】 ObjectiveTo construct single chain antibody from mAb SZ-21 against platelets GPⅢa.Methods Reverse transcription and polymerize chain reaction were used to clone the V region genes of mAb SZ-21(VH、VL).The cloned VH and VL were sequenced and constructed into a single chain antibody(ScFv).The fusion protein was expressed in E.coli .The activity of ScFv was characterized by ELISA and western blot.Results The VH and VL genes were homologous with the published gene sequences of mouse antibody variable region.Except for a little portein of secretory type,the recombinant protein was mostly in the form of inclusion bodies,and the yield was up to 21% of the total amount of bacteria protein.The ScFv fragment retained the binding activity to platelets.Conclusion Single chain antibody fragment of mAb SZ-21 was successfully constructed,which could bind specifically to platelets.The work may lay a foundation for application in clinical treatment in the fature. [更多还原