A Method for the Quantitative Assay of Inhibitor of Plasminogen Activation in Human Serum^[*]

Summary

A method for the assay of inhibitor of plasminogen activation in human serum, by means of a standard clot lysis system, is described.

1. A pure, stable inhibitor of activation, E. A. C. A., was used in the preparation of reference inhibitor curves.

2. The inhibitory activity of a serum specimen on plasminogen activation was expressed quantitatively in units arbitrarily equated with the millimolarity of an E. A. C. A. solution exerting an identical inhibitory effect.

3. The effects of time and temperature of storage of serum specimens, and of different methods of preparation of serum, on the results of the test have been determined. The use of different quantities of thrombin to clot the system did not alter the results.

4. No significant correlation was found between blood levels of inhibitor of plasminogen activation and antiplasmin in a group of 66 normal individuals. The temperature stability of the two types of inhibitor differed considerably, and the test system was significantly inhibited by the known activation inhibitor E. A. C. A. in concentrations too small to produce inhibition of a standard plasmin preparation. It was concluded that a discrete inhibitor of plasminogen activation was measured by this method.

5. The mean level of inhibitor of activation in a group of 66 normal subjects was 2.5 u (S. D. =0.5 u). There were no significant age or sex differences, and alimentary lipaemia was not found to cause significant changes.

^* This work was supported by a grant from the Tobacco Research Council of Great Britain on the recommendation of the British Heart Foundation.

^** Medical Research Council of Great Britain Junior Research Fellow.

• References

• 1 Norman P. S. Studies of the plasmin system. II. Inhibition of plasmin by serum or plasma. J. exp. Med 108: 53 1958;
• 2 Norman P. S, Hill B. M. Studies of the plasmin system. III. Physical properties of two plasmin inhibitors. J. exp. Med 108: 639 1958;
• 3 Sherry S, Lindemeyer R. I, Fletcher A. P, Alkjaersig N. Studies on enhanced fibrinolytic activity in man. J. clin. Invest 38: 810 1959;
• 4 Flute P. T. (1960) Proc. 7th Congr. europ. Soc. Haemat., Lond. 1959.
• 5 Paraskevas N, Nilsson I. M, Martinsson G. A method for determining serum inhibitors of plasminogen activation. Scand. J. clin. Lab. Invest 14: 138 1962;
• 6 McNicol G. P, Gale S. B, Douglas A. S. In-vitro and in-vivo studies of a preparation of urokinase. Brit. med. J I: 909 1963;
• 7 Dudok de Wit C. Investigations on the inhibitors of the fibrinolytic system. Thrombos. Diathes. haemorrh. (Stuttg.) 12: 105 1964;
• 8 Blix S. The quantitative determination of fibrinolytic inhibitors in plasma or serum by means of the fibrin plate. Scand. J. clin. Lab. Invest 16: 403 1964;
• 9 Bruce S. The effect of calcium on fibrinolysis in-vitro. J. clin. Path 17: 282 1964;
• 10 Ploug J, Kjeldgaard N. O. Urokinase, an activator of plasminogen in human urine. Biochem. biophys. Acta 24: 278 1957;
• 11 Alkjaersig N, Fletcher A. P, Sherry S. The activation of human plasminogen. I. Spontaneous activation in glycerol. J. biol. Chem 233: 86 1958;
• 12 Norman P. S. Studies of the plasmin system. I. Measurement of human and animal plasminogen. J. exp. Med 106: 423 1957;
• 13 Nilsson I. M, Olow B. Fibrinolysis induced by streptokinase in man. Acta chir. scand 123: 247 1962;
• 14 Alkjaersig N, Fletcher A. P, Sherry S. E-Aminocaproic acid: an inhibitor of plasminogen activation. J. biol. Chem 234: 832 1959;