P选择素表位结构、功能及其单克隆抗体抗粘附治疗研究

To further investigate the structure and function of P-selectin，the gene for lectin-EGF domain of P-selectin lectin-EGF was amplified from normal human platelets by RT-PCR，then was cloned into eukaryotic vector pEGFP-N1/lectin and pEGFP-N1/lectin-EGF. The recombinant plasmid was transfected into COS7 cells and was expressed in the cytoplasmic region. The prokaryotic vector pET42b(+)/lectin and pET42b(+)/lectin-EGF were transfected into E.coli BL 21 strain and expressed proteins were purified by affinity chromatography. Then the mAbs anti-lectin-EGF were prepared with classical hybridoma technique， and 3 hybridoma cell lines (B10，F3 and H5)were obtained with Ig subclasses, IgG2，IgG1 and IgG3 respectively， which their light chains were κ chain. The mAbs anti-lectin-EGF could specifically bind to fusion protein and recognize various antigenic determinants. Further investigation revealed that the mAbs could specifically recognize the natural P-selectin expressed on endothelial cell line stimulated by LPS，markedly inhibited adhesion between activated platelets and neutrophils. The findings of anti-adhesion/inflammation of mAbs would help to develop the new therapeutic agents and provide the basis of P-selectins 's structure and function.

用分子克隆技术进行P选择素基因在CHO细胞内表达，制备针对P选择素表位特异性的多种单寺】固澹鄄斓タ寺】固蹇拐掣郊岸陨鲈嗟谋；ぷ饔茫虿庑蚍治隹固寤颍煤怂幔被岱治鋈砑治鯬选择素表位结构与其功能的关系，筛选制备有效的抗粘附单克隆抗体，为抑制P选择素介导的早期炎症反应及肾小球肾炎特异性治疗提供有效的治疗手段。