Urokinase and non-urokinase fibrinolytic activity in protease-inhibitor-deprived plasma, assayed by a fibrin micro-plate method

Abstract

In the classical fibrin-plate assay, fibrinolytic activity is determined by measuring the area of the lysis zone formed when sample is applied on a planar fibrin gel. However, this method is characterized by low capacity and uncertainty in the determination of the lysis zones. To overcome these limitations an assay modified for microtiter plates was developed. Fibrin clots, with a suitable dye incorporated, were formed in wells of standard high adsorbtion microtiter plates. Each plate contained a serial dilution of urokinase as standard. Citrated test plasmas were treated with acetone to remove inhibitors before applied to the wells. The lyzate formed after appropriate incubation was removed, and the remaining volume of fibrin photometrically determined after being completely dissolved by plasmin. The fibrinolytic activity was determined as the difference in absorption before and after lysis. This is an accurate and relatively simple method for the assessment of urokinase and non-urokinase fibrinolytic activity in plasma. It is further a sensitive and quantitative fibrinolytic micro-technique with a high capacity.