The Rivanol Method for the Quantitative Determination of Antithrombin III in Plasma

Robert H. Yue; Toby Starr; Menard M. Gertler

doi:10.1055/s-0038-1649184

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1974; 31(03): 439-451
DOI: 10.1055/s-0038-1649184

Original Article

Schattauer GmbH

The Rivanol Method for the Quantitative Determination of Antithrombin III in Plasma

Robert H. Yue

¹Cardiovascular Research, Institute of Rehabilitation Medicine, New York University Medical Center, New York, New York 10016

,

Toby Starr

¹Cardiovascular Research, Institute of Rehabilitation Medicine, New York University Medical Center, New York, New York 10016

,

Menard M. Gertler

¹Cardiovascular Research, Institute of Rehabilitation Medicine, New York University Medical Center, New York, New York 10016

› Author Affiliations

Abstract

Summary

A simple quantitative method for the determination of antithrombin III in plasma is described. It is based on the preferential precipitation of plasma antithrombin activities by a chemical compound 2,5-diamino-7-ethoxyacridin lactate (rivanol). When the incubation time of plasma with rivanol took place within 0.5 and 2 hours under specifically controlled experimental conditions, rivanol precipitated α₂-macro-globulin selectively and quantitatively without any effect on antithrombin III. In the presence of rivanol, the inhibition of thrombin by plasma antithrombin III followed a second-order reaction with a rate constant of 80 1/M/sec. In this assay procedure, only the initial thrombin activity and the 30 minutes residual thrombin activity are required to evaluate the amount of antithrombin III present in the plasma quantitatively. “With proper routine laboratory care, this assay system was found to be accurate and reproducible.

Full Text

References

References
1 Abildgaard U. Purification of Two Progressive Antithrombin of Human Plasma. Scandinavian Journal of Clinical and Laboratory Investigation 1967; 19: 190
2 Abildgaard U. Binding of Thrombin to Antithrombin III. Scandinavian Journal of Clinical and Laboratory Investigation 1969; 24: 23
3 Abildgaard U, Egeberg O. Thrombin Inhibitory Activity of Fractions Obtained by Gel Filtration of Antithrombin III Deficient Plasma. Scandinavian Journal of Haematology 1968; 5: 155
4 Abildgaard U, Gravern K, Godal H. C. Assay of Progressive Antithrombin in Plasma. Thrombosis et Diathesis Haemorrhagica 1970; 24: 224
5 Bertho A, Grassmann W. Laboratory Methods of Biochemistry. New York: MacMillan & Co., Ltd.,; 1938
6 Biggs R, Denson K. W. E, Akman N, Borrett R, Hadden M. British Journal of Haematology 1970; 19: 283
7 Dombrose F. A, Seegers W. H, Sedensky J. A. Antithrombin: Inhibition of Thrombin and Autoprothrombin C (F — X_a) as a Mutual Depletion System. Thrombosis et Diathesis Haemorrhagica 1971; 26: 103
8 Heimburger N, Haupt H, Schwick H. G. Proteinase inhibitor of human plasma. In Fritz H, Tschesche H. (ed.) Proceedings of the International Research Conference on Proteinase Inhibitors, Munich, November 4—61970. 1971: 1
9 Heimburger N, Trobisch H. Blood Coagulation and Fibrinolysis. Angewandte Chemie International Edition 1971; 10: 85
10 Marciniak E, Tsukamura S. Two Progressive Inhibitors of Factor X_a in Human Blood. British Journal of Haematology 1972; 22: 341
11 Quick A. J. The Physiology and Pathology of Hemostasis . Philadelphia.: Lea and Febiger; 1951
12 Seegers W. H. Antithrombin as a Proteinase Inhibitor. Annals of New York Academy of Science 1968; 146: 593
13 Seegers W. H, Marciniak E. Inhibition of Autoprothrombin C Activity with Plasma. Nature 1962; 193: 1188
14 Seegers W. H, Smith H. P. Antithrombin Activity of Plasma: Quantitative Interrelationship. Proceedings of the Society of Experimental Biology, New York 1943; 52: 159
15 Seegers W. H, Cole E. R, Harmison C. R, Monkhouse F. C. Neutralization of Autoprothrombin C Activity with Antithrombin. Canadian Journal of Biology 1964; 42: 359
16 Steinbuch M, Beatrix C, Josso F. A₂-macroglobulin as a Progressive Antithrombin. Nature 1967; 216: 500
17 Steinbuch M, Quentin M, Pejaudier L. Specific Technique for the Isolation of Human A₂-macroglobulin. Nature 1965; 205: 1227
18 Yin E. T, Wessler S, Butler J. V. Plasma Heparin: A Unique, Practical, Submicrogram-Sensitive Assay. Journal of Laboratory and Clinical Medicine 1973; 81: 298
19 Yin E. T, Wessler S, Stoll P. J. Rabbit Plasma Inhibitor of the Activated Species of Blood Coagulation Factor X. Purification and Some Properties. Journal of Biological Chemistry 1971; a 246: 3694
20 Yin E. T, Wessler S, Stoll P. J. Biological Properties of the Natural Occurring Plasma Inhibitor to Activated Factor X. Journal of Biological Chemistry 1971; b 246: 3703
21 Yin E. T, Wessler S, Stoll P. J. Identity of Plasma-Activated Factor X Inhibitor with Antithrombin III and Heparin Cofactor. Journal of Biological Chemistry 1971; c 246: 3712
22 Yue R. H, Starr T, Gertler M. M. Antithrombin III Levels in Ischemic Thrombotic Vascular Diseases. Proceedings of the III Congress of the International Society of Thrombosis and Haemostasis, August 22–261972. Washington, D. C.: 1972
23 Yue R. H, Starr T, Gertler M. M. The Rivanol Method for the Quantitative Determination of Antithrombin III in Plasma. Circulation 1973; a 48 Supplement 77
24 Yue R. H, Starr T, Gertler M. M. Quantitative Determination of Total Antithrombin III in Plasma. Thrombosis et Diathesis Haemorrhagica 1973; b 30: 84