We have studied the effects of PAI-1 on the conversion of scu-PA into tcu-PA in vitro in plasma containing or not a 125I-fibrin clot by determining tcu-PA activity on S2444. Two preparations of PAI-1 have been used, a fraction of medium conditioned with the monkey Vero cells (Vero-Prep), the antiurokinase activity of which is inhibited at 83% by anti PAI-1 IgG, or purified human PAI-1 from HT 1080 fibrosarcoma cells. Scu-PA purified from human kidney cells has been treated with diisopropylfluorophosphate before use. In plasma, conversion of scu-PA into the tc form is accelerated by addition of anti PAI-1 IgG. In plasma containing a clot, generation of tcu-PA, is considerably delayed after addition of the Vero-Prep or human PAI-1. Clot lysis is also decreased but to a lesser extent than it would be expected from the level of tcu-PA activity. Addition of anti PAI-1 antibodies shortens the lag phase before tcu-PA appears and moderatly accelerates clot lysis. These results demonstrate the importance of PAI-1 for the stability of scu-PA in plasma in vitro by delaying its conversion into tcu-PA.