Sensitive fluorometric determination of plasminogen activator in cell lysates and supernatants

A fluorogenic substrate for plasmin, CBZ-Gly-Pro-Arg-AEC, has been synthesized and used to develop a new sensitive photometric and fluorometric assay of plasminogen activator activity. The fluorescence intensity of free AEC at 460 nm is about 3 orders of magnitude higher than that of acyl-AEC. The release of AEC from the peptidyl derivative was monitored fluorometrically after extraction of free AEC in ethylacetate. Under such conditions, the Km was 0.16 mM. This method was used to monitor the activity of plasminogen activator synthetized by fibroblastic cells (BHK 21 C 13) either released in the supernatants or cell-associated.