单克隆抗体亲和层析法纯化人尿激酶

Purification of Human Urokinase byMonclonal Antibody Affinity Chromatography

含抗人尿激酶单克隆抗体的腹水经ＰｒｏｔｅｉｎＡ－ＳｅｐｈａｒｏｓｅＣＬ４Ｂ柱纯化得到ＩｇＧ纯品，分子量为１５０ｋＤ（重链５５ｋＤ，轻链２０ｋＤ），与ＣＮＢｒ－Ｓｅｐｈａｒｏｓｅ４Ｂ偶联制成抗人尿激酶单克隆抗－体－Ｓｅｐｈａｒｏｓｅ４Ｂ亲和层析介质。尿激酶粗品经单抗－Ｓｅｐｈａｒｏｓｅ４Ｂ亲和层析柱纯化后，所得尿激酶制品比活为９×１０－４～１．７×１０－３ｍｏｌ／ｓ·ｍｇ－１，纯化倍数为１５～３０倍，酶活性回收率在５０％以上。

Purified monoclonal antibodies against human urokinase by Protein A-Sepharose CL 48 are coupled to CNBr-Sepharose 4B. SDS-PAGE of the purified urokinase from crude urokinase shows a single band (54000 daltons) Over 50% of enzyme activity is recovered from the column The specific activity is 9×10-4～1. 7×10-3 mol/s . mg-1, attaining a purififcation factor of 15 to 30. With a single-step procedure active urokinase from crude preparations is obtained.