人红细胞凝集型单抗的制备与鉴定

目的制备人红细胞凝集型单克隆抗体并进行鉴定。方法以3种方式处理人血红细胞,获得红细胞免疫原,分别免疫Balb/c小鼠。应用杂交瘤技术,取免疫小鼠的脾细胞与Sp2/0骨髓瘤细胞融合,以凝集实验筛选分泌凝集型抗体的杂交瘤细胞,继而用有限稀释法克隆3次,并对杂交瘤细胞的稳定性进行测定。通过腹水法制备抗体,亲和层析法纯化抗体,对抗体的特性进行研究。结果 3种方式获得的免疫原均能建立有效的免疫反应,低渗加超声破碎法所获得的小鼠血清抗体效价最高。通过融合获得8株可稳定分泌抗体的杂交瘤细胞,其中,4A1分泌的抗体凝集效价最高。抗体可使4种人血型的红细胞凝集,无种属交叉凝集反应。结论有效制备了针对人红细胞的凝集性单抗,为其之后的应用研究奠定了基础。福建省省属公益类项目(2015R1036-3,2016R1034-2,2017R1036-3);;福建省科技计划(2014Y0082,2014Y0083

Construction of a Neotype Controllability Magnetic Targeting Restructuring Truncated Tissue Factor

目的:构建新型的具有可控性且能诱发动物血栓的栓塞剂磁靶向凝血蛋白,从而为动物血栓模型的建立提供新工具。方法:使用化学共沉淀法合成磁流体(; magnetic nanoparticles, MNPs),通过透析对其纯化后用超导量子干涉磁强针(; SQUID)测定其磁性能;利用基因工程方法表达截短组织因子( truncated tissue factor,tTF); (又称为凝血蛋白)并纯化与鉴定;采用戊二醛交联法构建磁靶向凝血蛋白(; MNPs-tTF),体外分析其磁控靶向性及凝血效力。结果:成功构建了磁靶向凝血蛋白MNPs-tTF,体外实验证实其保留有磁流体MNPs的磁控靶向; 性,以及凝血蛋白tTF的凝血效力。结论:成功构建了磁靶向凝血蛋白MNPs-tTF,为血栓模型动物的建立提供一种新工具。Objective: To establish a neotype controllability magnetic targeting; restructuring truncated tissue factor s for establish animal models of; thrombosis. Methods: Magnetic nanoparticles ( MNPs) were prepared by; chemical coprecipitation,and employed the superconducting quantum; interference device to characterized the MNPs magnetization. The; truncated tissue factor ( tTF) were expressed efficiently in E. coli and; purification by gene engineering technology. MNPs conjugated the tTF by; glutaraldehyde to compose MNPs-tTF. In vitro analysis the effects of; magnetic targeting and blood coagulation of MNPs-tTF. Result: The; magnetic targeting restructuring truncated tissue factor composites were; build successfully. They are reserve efficient coagulation effect and; magnetic targeted ability in vitro. Conclusion: We have successfully; prepared magnetic targeting thrombin system MNPs-tTF have been; successfully prepared,which maybe provide a basis for developing the; establish animal models of thrombosis.福建省科技计划重点项目; 福建省属公益类项