摘要
本文介绍了一种简便易行的纯化组织型纤溶酶原激活剂(t-PA)的方法。新鲜猪心组织经丙酮脱脂脱水,制成干粉,再经醋酸钾缓冲液提取。提取液经阳离子交换柱、肝素柱亲和层析及凝胶柱层析分离纯化。经SDS凝胶电泳分析证明纯化的t-PA只在分子量为67000道尔顿位置出现一条染色带。同时把该蛋白带转移到纤维蛋白板上,也在同一位置出现溶解带。
A simplified procedure for the purification of porcine heart tissue-type plasminogen activator (t-PA) was described.Fresh porcine hearts were delipidated by acetone treatment and dried.The delipidated powder was extracted with potassium acetate.The extract was fractionated with ammonium sulfate, and purified by combination of cation-exchange chromatography on SP-sephadex C-50, affinity chromatography on heparin-spharose 4B and gel filtration on sephadex G-150.The final product gave a single protein band (apparent molecular weight.67000) on SDS-PAGE and on fibrin autography of t-PA by transfering into fibrin agar gel.The activity of purified t-PA was completely inhibited by specific anti-t-PA antibody and the lysis zone had not appeared on the fibrin-agar plate without plasminogen.
关键词
纯化
鉴定
纤溶酶激活剂
T-PA
Tissure-type plasminogen activator
SDS-PAGE
Fibrin autography on fibrin-agar gel
Heparin-sepharose 4B column
