Detergent tween 80 modifies the specific activity of PAI-1

Abstract

Summary In the measurement of the reaction between low concentrations of tissue plasminogen activator (tPA) and plasminogen activator inhibitor type-1 (PAI-1) in buffer, a detergent is needed to optimize the assay and to prevent unspecific adhesion. Not only the recovery, but also the activity of tPA and PAI-1 depends on the type of detergent and its concentration. Stable values for the specific activities were obtained in the concentration range of 0.03–0.3 g/L Tween. Moreover, it became apparent that the composition of Tween also seems to affect PAI-1, since the specific activity of PAI-1 in a buffer with Tween 80 from one source was approximately 40–50% lower than with Tween 80 from another source. SDS-PAGE confirmed the formation of a correspondingly smaller amount of stable complex with tPA, but also the concomitant appearance of cleaved, ‘substrate’ PAI-1. The observed decrease in specific activity in the reaction with tPA thus depends on an increase in the relative amount of cleaved PAI-1. In contrast, the same high specific activity was consistently obtained with Tween 20 purchased from different sources. The difference between the different commercial preparations of Tween 80 with regard to the properties of PAI-1 was also found in a clot lysis assay in which thrombin, fibrinogen and plasminogen (the natural substrate for tPA) were used. Although we have not been able to identify a specific component in Tween, our observations can be of practical use for other workers in this field.