AN ENZYME-IMMUNOASSAY FOR THE SIMULTANEOUS DETERMINATION OF ACTIVE TYPE-1 PLASMINOGEN-ACTIVATOR INHIBITOR (PAI-1), AND T-PA PAI-1 COMPLEXES/

An assay has been developed which enables the simultaneous determinati on of active free PAI-I and t-PA/PAI-1 complexes in plasma samples. Te st plasma is divided into two subsamples in which (a) active PAI-I is quantitatively converted to t-PA/PAI-1 complexes, and (b) conversion o f PAI-l to t-PA/PAI-1 complexes is prevented. The amounts of t-PA/PAI- 1 complex in the two subsamples are then assessed in a one-step enzyme immunoassay (EIA) consisting of a monoclonal anti-PA antibody, and a HRP conjugate of polyclonal anti-PAI-l antibodies. The amounts of t-PA /PAI-1 complexes in the two subsamples are the t-PA/PAI-1 complex conc entrations at the time of sampling (b); whereas the difference between the two values (a)-(b) equals the amount of active PAI-1. The assay h as a high sensitivity, specificity, accuracy and precision. The time-t o-result is 45 h. It meets the criteria set by the Scientific and Stan dardization Committee (SSC) of the International Society of Thrombosis and Haemostasis (ISTH) for a candidate reference method.