单克隆抗体制备的亲和填料在血浆高丰度蛋白分离中的应用  

A Study of Monoclonal Antibody Prepared Immunoaffinity Resin for Separating High Abundant Proteins in Human Plasma Samples

作　　者：王云丹[1] 宁云山[1] 彭丹丹[2] 李妍[1] 方勤美[3] 邓新宇[3] 姜颖[3] 李明[1] 

机构地区：[1]南方医科大学生物技术学院,广州510515 [2]南方医科大学附属珠江医院心内科,510282 [3]军事医学科学院放射医学研究所,北京100850

出　　处：《中国生物工程杂志》2008年第5期71-77,共7页China Biotechnology

基　　金：国家“863”计划(2006AA02A311);国家“973”计划(2001CB5102)资助项目

摘　　要：分离去除高丰度蛋白成为血浆蛋白质组研究的关键问题之一。从已建株的17株抗人血浆白蛋白单克隆抗体库中选出一株合适抗体。该抗体用Protein G Sepharose 4B吸附,然后用交联剂DMP(庚二亚氨酸二甲酯二盐酸盐)使之与Protein G蛋白共价交联。实验结果显示600μl(结合1mg抗体)胶能分离10μl血浆中的白蛋白和球蛋白。填料重复使用10次后吸附能力无显著改变。对填料的吸附蛋白进行MALDI-TOF/TOF鉴定主要为白蛋白和球蛋白及其碎片,但发现吸附的蛋白有转铁蛋白,纤维蛋白原,抗胰蛋白酶,前脂蛋白,Vitamin D结合蛋白,Keratin 10和CD5抗原相似蛋白。实验结果表明该方法能在固定目的抗体的同时最大量地保持抗体活性,该方法有望在蛋白质组学研究的高丰度蛋白分离中得到广泛应用。High abundant plasma proteins deteriorated the separating results of plasma proteins in plasma proteomics research ,it is necessary to separate high abundant proteins in plasma samples. Protein G was used to absorb the anti-albumin monoclonal antibodies then cross4inked with mAb by Dimethyl pimelinediimidate chloride. The results suggested that 10007 g antibody was site-directed immobilized and kept the capacity of binding around 5009. g albumin proteins after 10 cycles of elution-regeneration-binding. The bound proteins included albumin, IgG, vitamin D binding protein,Fibrinogen,Transthyretin, Keratin 10, Transferrin, C3, proapolipoprotein, CD5 antigen-like protein and alphal - antitrypsin. A prevailing,applicable and valuable approach was shomed to separate high abundant proteins in human plasma as well as other samples.

关 键 词：单克隆抗体 定向固定 高丰度蛋白 血浆蛋白质组 

分 类 号：Q819[生物学—生物工程]