摘要
目的 在HeLa细胞中表达抗人γ 精浆蛋白 (γ Sm)单链抗体 (scFv)基因 ,并进行亲和活性测定。方法 在已经构建抗人γ SmscFv基因基础上 ,将基因克隆入真核分泌表达载体 pSecTag2 B中 ,并转染HeLa细胞进行表达。表达产物纯化后 ,用流式细胞仪进行亲和活性测定。结果 表达产物经SDS PAGE和Western印迹实验证实为约30ku的特异蛋白条带 ,纯化后经流式细胞仪检测可以特异性地结合PC 3细胞。结论 获得了可与PC 3细胞特异结合的抗人γ SmscFv 。
Objective To express anti-human γ-seminoprotein (γ-Sm) single chain Fv antibody (scFv) gene in HeLa cells and identify its binding affinity for PC-3 cells. Methods The anti-human γ-Sm scFv was cloned into the pSecTag2-B expression plasmid. Then the pSecTag2-B plasmids containing the anti-human γ-Sm scFv gene were transfected by HeLa cells. The expression products were analyzed by both SDS-PAGE and Western blot, then were purified with Ni 2+-NTA superflow affinity chromatography. The binding affinity for PC-3 cells was measured by flow cytometry. Results The expression products of the anti-human γ-Sm scFv, whose relative molecular mass (Mr) was about 30 000, were confirmed by SDS-PAGE and Western blot. After being purified with Ni 2+-NTA superflow affinity chromatography, the anti-human γ-Sm scFv showed significantly strong binding with PC-3 cells. Conclusion The anti-human γ-Sm scFv which can bind with PC-3 cells has been successfully obtained for the potential use in clinical studies.
出处
《西安交通大学学报（医学版）》
CAS
CSCD
北大核心
2003年第5期420-422,共3页
Journal of Xi’an Jiaotong University（Medical Sciences）
基金
国家自然科学基金 (No .3990 0 1 80 )
全军重点实验室研究基金 (No .1 997 71 2 2 )
