Immunological Assay for the Determination of Procarboxypeptidase U Antigen Levels in Human Plasma

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Abstract
The importance of carboxypeptidase U as a novel regulator of the fibrinolytic rate has attracted a lot of interest recently. In the present work, an ELISA was developed using polyclonal antibodies raised against recombinant proCPU, expressed in DON cells. The assay determines the antigen concentration of the zymogen of carboxypeptidase U, procarboxypeptidase U, in human citrated plasma or EDTA plasma. No interference is observed with plasma carboxypeptidase N. The assay is very reproducible (within-run: 4.6% CV, between-run: 6.8% CV). In a group of 479 healthy individuals the mean proCPU antigen concentration is 13.4 μg/ml (SD 2.5 μg/ml). A good correlation is found with the functional procarboxypeptidase U assay described earlier (r = 0.82, p <0.0001) (Schatteman K, Goossens F, Scharpé S, Neels H, Hendriks D Clin Chem 1999; 45: 807-813). The significant correlation between the proCPU antigen concentration and the 50% clot lysis time stresses its importance as a player in fibrinolysis control. Abbreviations: CPU, carboxypeptidase U; proCPU, procarboxypeptidase U; TAFI, thrombin activatable fibrinolysis inhibitor; AP, alkaline phosphatase; EDTA, ethylenediamine tetra-acetic acid; Tris, tris-(hydroxyethyl)-aminomethane; PPACK, phenylalanyl-prolyl-chloromethyl ketone; SP, sulphopropyl; HEPES, N-2-hydroxyethylpiperazine-N’-ethanesulfonic acid