Electrochemical assay of plasminogen activators in plasma using polyion-sensitive membrane electrode detection

Two relatively simple electrochemical assay methods suitable for the measur ement of plasminogen activators (including urokinase (u-PA), streptokinase (SR), and tissue plasminogen activator (t-PA)) in plasma samples are descri bed. In one approach, the initial rate of decrease in the potentiometric re sponse of a polycation-sensitive membrane electrode toward protamine is mon itored after addition of a preincubated reaction mixture containing the sam ple and exogenous plasminogen. The plasmin formed from plasminogen by the a ctivators catalyzes the decomposition of the arginine-rich protamine substr ate, yielding smaller polycationic fragments that are not sensed by the ele ctrode. Alternately, the sample, plasminogen, and protamine can be incubate d together, and the remaining protamine in this reaction mixture can be mea sured at a fixed point in time by placing the electrode into the mixture an d recording the electromotive force response. Working curves found with bot h methods for plasma samples spiked with varying levels of the activators c over the expected therapeutic activity ranges found in the plasma of patien ts treated with these "clot-busting" drugs. (C) 1999 Academic Press.