Sensitive and Direct Assays for Functionally Active Plasminogen Activators (tissue-type and urokinase-type) in Plasma

Abstract

A previously described bioimmunoassay for tissue-type plasminogen activator (t-PA) has been modified in terms of antibody concentration and conditions of incubation to achieve a sensitivity range of approximately 5-500 IU/L of t-PA. A similar assay has been developed for urinary-type plasminogen activator (U-PA), also known as urokinase (UK), achieving a sensitivity range of approximately 5-500 X 10(-1) IU/L. The direct sensitive t-PA assay has been demonstrated to be a quantitative alternative to the traditional semiquantitative euglobulin clot lysis time (ECLT) assay, with correlation coefficients of 0.967 and 0.914, depending on the laboratory where the ECLT was carried out. This assay has the added advantages of avoiding loss of t-PA or UK during the formation of the euglobulin fraction and of calibration in terms of International Standards for both t-PA and UK. With both these assays, it has been demonstrated that the enhanced fibrinolytic activity observed after exercise results from increased levels of t-PA while the level of UK in plasma remains unaltered. Free t-PA (10-60 IU/L) has been demonstrated in resting plasma, whereas the level of free UK activity was 50-100 IU/L. Venous occlusion of matched groups of subjects with and without deep venous thrombosis (DVT) showed a wide variation of response in t-PA levels in each group and no change in UK levels. There was no difference between the DVT and non-DVT groups, and this suggests that the notion of the "nonresponder" being more prone to thrombosis needs to be reexamined.

The tissue plasminogen activator and urokinase response in vivo during natural resolution of venous thrombus

“…The t-PA and urokinase concentrations were measured with a bioimmunoassay, by use of affinity purified polyclonal bovine anti-human recombinant t-PA (Welcome Biotechnology, Beckenham, United Kingdom) and rabbit anti-human u-PA (National Institute for Biological Standards & Controls, South Mimms, United Kingdom) antibodies, respectively, for the immunocapture phase. 23 The detection buffer was composed of LYS-plasminogen and the chromogenie substrate S-225 1 (Chromogenics, M61ndal, Sweden) in tris-buffered saline solution. Standard dilution curves with the International Standards for t-PA (87 of 670) and u-PA (87 of 594) were used.…”

“…Although urokinase-type plasminogen activator (u-PA) is known to be produced by vascular endo-From the Department of Surgery, St. Thomas' Hospital, and Imperial Cancer Research Fund (Mr. Soo and Dr. Bobrow), Immunohistology Laboratory, London, and the National Institute of Biological Standards and Control (Dr. Gaffney), South Mimms. Presented at the Seventh Annual Meeting of the American Venous Forum, Fort Lauderdale, Fla., Feb. [23][24][25]1995. Reprint requests: Andrew D. R. Northeast, M.A, FRCS, Department of Surgery, St. Thomas' Hospital, Lambeth Palace Road, London SE17EH United Kingdom.…”

Monoclonal antibodies to crosslinked fibrin degradation products (XL‐FDP) II. EVALUATION IN A VARIETY OF CLINICAL CONDITIONS

“…The procedure here described is a modified version of that reported elsewhere with these mabs (Gaffney et al, 1987). However, details of the mabs used in this assay (their preparation, purification and labelling with horseradish 50 -20-peroxidase) and the method of preparation of the antigen for use as a standard for X-oligomer are already described elsewhere (Gaffney et al, . 1987.…”

Aprotinin reduces cardiopulmonary bypass‐induced blood loss and inhibits fibrinolysis without influencing platelets

“…PFA was determined by the method of Mahmoud-Alexandroni et a1 (1989) which measures the activity of free plasma tPA. Data obtained with this assay correlate with euglobulin clot lysis times (Mahmoud-Alexandroni et al, 1989).…”