A fluorescent substrate assay for plasminogen

Pochron, Sharon P.; Mitchell, Gary; Albareda, Ileana; Huseby, Rolf M.; Gargiulo, Robert J.

doi:10.1016/0049-3848(78)90179-2

Cited by 35 publications

(8 citation statements)

References 11 publications

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“…Harris et al 27 compared the automated CBZlys-SBZ substrate test for plasminogen with a fluorogenic substrate assay. 56 The correlation coefficient 26 and Teger-Nilsson 69 re ported an automated α 2 -PI synthetic substrate assay using S-2251 and an LKB 8600 reaction rate analyzer modified to dispense two reagents simul taneously every 30 seconds. The incubation time between plasmin and the patient sample was set at 30 seconds to avoid the detection of inhibitors other than α 2 -PI.…”

Section: Assays For the Fibrinolytic Systemmentioning

confidence: 99%

Development and Clinical Use of Automated Synthetic Substrate Methods for the Evaluation of Coagulation and Fibrinolysis

Mitchell¹

1983

Semin Thromb Hemost

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Section: Assays For the Fibrinolytic Systemmentioning

confidence: 99%

Development and Clinical Use of Automated Synthetic Substrate Methods for the Evaluation of Coagulation and Fibrinolysis

Mitchell¹

1983

Semin Thromb Hemost

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“…Another assay method for plasminogen was described in which instead of a chromo genic substrate a fluorescent substrate was used (15). With the exception of the sub strate the procedure is approximately the same as the method described by (6) and the results are also identical.…”

mentioning

confidence: 59%

Basics and Practice in Evaluating Plasminogen

Vinazzer¹

1988

Pathophysiol Haemos Thromb

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There exist different ways of assays of plasminogen which give information about different properties of this proenzyme. The concentration of plasminogen can be determined by its antigenicity. Since the normal concentration of plasminogen in plasma is between 15 and 25 mg/dl the test can be carried out by simple methods such as radial immunodiffusion on Partigen® plates. The possibility of errors is small and there is no need of special apparatus. The disadvantages are the lapse of 24 h until the result is available and the fact that the knowledge of the concentration does not give any information about the activity. The activity can be measured by different coagulation tests. A typical assay would involve activation of plasminogen to plasmin, addition of plasminogen-free thrombin and measuring of the lysis time. The result is however, dependent on more than one variable. Plasmin is rapidly inhibited by alpha-2-antiplasmin (APL) and there is also a dependence of the lysis time on the amount of clottable fibrinogen in the test system. Better results can be obtained by the use of diluted test plasma and addition of a constant amount of plasminogen-free fibrinogen. A different way would be the use of the euglobulin fraction instead of plasma. This has however, the possible disadvantage of incomplete precipitation of plasminogen. Instead of coagulation tests the activity can also be determined when diluted activated plasma is placed on plasminogen-free fibrin plates and the amount of lysis in the plate is recorded. All assays of this group also depend on the method of activation of plasminogen. There may be different results when tests are simultaneously carried out after activation with streptokinase, urokinase and t-PA respectively. A different way of determining plasminogen activity is the use of chromogenic or fluorogenic substrates. These methods have different advantages. Some substrates are highly sensitive to the SK-plasminogen complex which is formed in plasma when SK is added in considerable abundance. This complex is not inhibited by APL and there is no fibrin involved in this type of reaction. The method is, therefore, more specific than most coagulation tests. The splitting of the substrate is measured photometrically as an endpoint test or as a kinetic reaction. For clinical purposes the determination of plasminogen activity by a chromogenic substrate method is sufficient. Only in order to detect abnormal plasminogen molecules an additional test of plasminogen concentration is a minimum requirement which is then supplemented by attemps of activation with different substances such as urokinase and t-PA. Plasminogen assays in clinical practice are used for different reasons. Congenital deficiency or a defective plasminogen molecule with increased thrombotic tendency have been described but are very rare. Acquired deficiency of plasminogen is found in severe liver dysfunction, in DIC and during therapy with SK or UK. In the latter cases assays of plasminogen activity are or greater clinical importance and ...

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“…Pochron et al [357] recently reported using the synthetic oligopeptide sub strate D-valyl-L-leucyl-L-lysyl-5-amidoisophthalic acid dimethylester for plas min by fluorescent detection. Plasma plasminogen was assayed fluorometrically following conversion of plasminogen to plasmin using streptokinase for enzyme activation.…”

Section: Chemical Namementioning

confidence: 99%

“…Friberger and Knos extended the use of the p-nitroaniline substrate, S 2251, to the study of plasminogen in human plasma [133]. A mean value of approximately four caseinolytic units per ml (CTA U/ml) of plasma were found in the normal populations; however, Pochron et al [357] showed a normal range within populations of approxi mately 2.2 to 4.0 CTA U/ml. Soria and Samama [428] recently reported on the use of D-Val-Leu-Lys-pnitroanilide, S 2251, the substrate previously described by Friberger, by fol lowing patients undergoing thrombolytic therapy and comparing parallel levels of plasminogen and fibrinogen.…”

Section: Chemical Namementioning

confidence: 99%