Thromb Haemost 1977; 38(02): 0494-0503
DOI: 10.1055/s-0038-1651855
Original Article
Schattauer GmbH

The Different Forms of Antithrombin III in Serum[*]

D. S Pepper
1   S. E. Scotland Regional Blood Transfusion Centre, Royal Infirmary, Edinburgh, Scotland
,
D Banhegyi**
1   S. E. Scotland Regional Blood Transfusion Centre, Royal Infirmary, Edinburgh, Scotland
,
J. D Cash
1   S. E. Scotland Regional Blood Transfusion Centre, Royal Infirmary, Edinburgh, Scotland
› Author Affiliations
Further Information

Publication History

Received 12 October 1976

Accepted 04 April 1977

Publication Date:
04 July 2018 (online)

Summary

Antithrombin III (AT III) complexes were isolated from human serum by affinity chromatography and gel filtration. In the first step of the preparation, using heparin-agarose chromatography, we observed that the complexed form of AT III bound less strongly to the gel than the free form and that about half of the AT III was free. With further purification a 2.5 × 105 molecular weight complex was isolated. Using 125I labelled human thrombin, this complex was radioactive indicating the presence of thrombin. Only in a synthetic thrombin-AT III system was a 9 × 104 molecular weight complex detected, but not in serum. These facts suggest that in serum AT III complexes may exist in a polymeric form. Also, an AT III antigen derived from the original AT III molecule, but not complexed, was isolated which may be a degradation product.

Abbreviations used: AT-III, antithrombin III. Hepes, N-2-Hydroxyethylpiperazine-N-2-Ethanesulphonic acid.

* A preliminary account of this work was presented at the one day Scientific meeting of the British Society of Haematology, held at the Middlesex Hospital, London, in January, 1976 (Pepper and Banhegyi, 1976).


** Present address: 1st Department of Medicine, Postgraduate Medical School, Budapest, Hungary. Correspondence and reprint requests to D. S. Pepper.


 
  • References

  • 1 Abildgaard U. 1967; Purification of two progressive antithrombins of human plasma. Scandinavian Journal of Clinical and Laboratory Investigation 19: 190.
  • 2 Abildgaard U. 1969; Binding of thrombin to antithrombin III. Scandinavian Journal of Clinical and Laboratory Investigation 24: 23.
  • 3 Binder B. 1973; On the complex formation of antithrombin III with thrombin. Thrombosis et Diathesis Haemorrhagica 30: 280.
  • 4 Damus P. S, Hicks M, Rosenberg R. D. 1973; Anticoagulant action of heparin. Nature 246: 355.
  • 5 Damus P. S, Wallace G. A. 1974; Purification of canine antithrombin Ill-heparin cofactor using affinity chromatography. Biochemical and Biophysical Research Communications 61: 1147.
  • 6 Daussant J, Skakoun A. 1975; Combination of pore gradient polyacrylamide gel electrophoresis and crossed agarose gel Immunoelectrophoresis. Journal of Immunological Methods 7: 39.
  • 7 Godal H. C. 1961; The assay of heparin in thrombin systems. Scandinavian Journal of Clinical and Laboratory Investigation 13: 153.
  • 8 Hunter W. M. 1971. In: Kirkham K. E, Hunter W. M. Radioimmunoassay Methods. 3 Churchill-Livingstone; Edinburgh.:
  • 9 Lanchantin G. F, Plesset M. L, Friedmann J. A, Hart D. W. 1966; Dissociation of esterolytic and clotting activities of thrombin by trypsin-binding macroglobulin. Proceedings of the Society for Experimental Biology and Medicine 121: 1184.
  • 10 Machovich R, Blasko G. Y, Himer A, Szikla K. 1975; Effect of sodium and potassium ions on the activity of human antithrombin-heparin cofactor. Thrombosis Research 7: 305.
  • 11 Marciniak E. 1975; Adverse effect of heparin in thrombin-antithrombin III interaction. Thrombosis et Diathesis Haemorrhagica 34: 748.
  • 12 Middleton S. M, Bennet J. H, Smith J. K. 1973; A therapeutic concentrate of coagulation factors II, IX and X from citrated factor VIII depleted plasma. Vox Sanguinis 24: 441.
  • 13 Miller-Andersson M, Borg H, Andersson L.-O. 1974; Purification of antithrombin III by affinity chromatography. Thrombosis Research 5: 439.
  • 14 Pepper D. S, Banhegyi D. 1976; Studies on complexes of antithrombin III produced in serum and from artificial mixtures of activated coagulation factors. British Journal of Haematology 33: 146.
  • 15 Porath J, Aspberg K, Drevin H, Axen R. 1973; Preparation of cyanogen bromide-activated agarose gels. Journal of Chromatography 86: 53.
  • 16 Rosenberg R. D, Damus P. S. 1973; The purification and mechanism of action of human antithrombin-heparin cofactor. Journal of Biological Chemistry 248: 6490.
  • 17 Rosenberg R. D. 1974; The effect of heparin on factor XIa and plasmin. Thrombosis et Diathesis Haemorrhagica 33: 51.
  • 18 Sas G, Pepper D. S, Cash J. D. 1975; Investigations on antithrombin III in normal plasma and serum. British Journal of Haematology 30: 265.
  • 19 Thaler E, Schmer G. 1975; A simple two-step isolation procedure for human and bovine antithrombin II/III (Heparin cofactor) a comparison of two methods. British Journal of Haematology 31: 233.
  • 20 Vennerod A. M, Laake K. 1975; Inhibition of purified plasma kallikrein by antithrombin III and heparin. Thrombosis Research 7: 223.
  • 21 Wessler S, Reimer S. M, Sheps M. C. 1959; Biologic assay of a thrombosis-inducing activity in human serum. Journal of Applied Physiology 14: 943.
  • 22 Yin E. T. 1974; Effect of heparin on the neutralization of Factor Xa and thrombin by the plasma alpha-2-globulin inhibitor. Thrombosis et Diathesis Haemorrhagica 33: 43.