单克隆抗体SZ-95亲和色谱纯化人血小板第4因子

目的用单克隆抗体亲和色谱从人血小板破碎液中纯化血小板第 4因子 (PF4 )。方法将单克隆抗体SZ 95 IgG与溴化氰活化的Sepharose 4B凝胶连接成亲和色谱柱SZ 95 IgG Sepharose 4B ,人血小板破碎液经此亲和色谱柱上样后 ,经洗脱获得PF4 ,采用 15 %SDS 聚丙烯酰胺凝胶电泳鉴定其纯度 ,用点印迹鉴定其免疫活性。结果SZ 95 Sepharose 4B亲和色谱柱的偶联率为 72 % ,每 1ml(约 1× 10 9个血小板 )血小板破碎液中可以纯化到PF4 18μg ,其相对分子量约为 12kD ,经点印迹显示与单抗SZ 95反应显带。结论用SZ 95 Sepharose 4B亲和色谱柱纯化的PF4产品得率高、纯度高、活性好

Purification of human platelet factor 4 by monoclonal antibody (SZ-95) affinity chromatography

PurposeThe aim is to purify human platelet factor 4 (PF4) using a monoclonal antibody affinity chromatography.MethodsAfter binding a monoclonal antibody against PF4 (SZ 95) to CNBr activated sepharose 4B, a SZ 95 sepharose 4B affinity column was got. Proteins, which were purified from triton X 100 solubilized platelet solution by SZ 95 sepharose 4B affinity chromatography, were characterized by 15% SDS PAGE and Dot blot.ResultsThe binding rate of SZ 95 to sepharose 4B is 72% and 18 μg of PF4 can be purified from 1×10 9 of platelets. SDS PAGE and Dot blot indicated that this 12kD protein was PF4.ConclusionUsing SZ 95 sepharose 4B affinity chromatography to purify PF4 is an efficient method to prepare PF4 with high biological activity.