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Thromb Haemost 1981; 46(04): 699-705
DOI: 10.1055/s-0038-1653456
Original Article
Schattauer GmbH Stuttgart

Rabbit Antibodies Against the Procoagulant Activity (VIII:C) of Human Factor VIII

Some Theoretical and Practical Considerations on the Human Factor VIII Molecule Using Heterologous Antibodies
T H Tran
The Coagulation and Fibrinolysis Laboratory, Kantonsspital, Basle, Switzerland
,
G A Marbet
The Coagulation and Fibrinolysis Laboratory, Kantonsspital, Basle, Switzerland
,
F Duckert
The Coagulation and Fibrinolysis Laboratory, Kantonsspital, Basle, Switzerland
› Author Affiliations
Further Information

Publication History

Received 05 August 1981

Accepted 23 September 1981

Publication Date:
05 July 2018 (online)

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Summary

The procoagulant activity VIII:C was separated from factor VIII antigen (VIIIR:Ag) by gel filtration in the presence of 0.25 mol/l calcium chloride. Antibodies (anti-VIII:C) were obtained by immunization of rabbits with VIII:C. The last step of the purification procedure of antibodies consists of an adsorption on VIIIR:Ag-Sepharose 2 BCL as immunoadsorbent to remove contaminating traces of antibodies against VIIIR:Ag. The anti- VIII:C titer remains unchanged during this adsorption (29 Bethesda units per mg). In solution, anti-VIII:C neutralies factor VIII activity (in plasma, cryoprecipitate or in purified form) and the fragment VIII:C without reacting with VIIIR:Ag. Once immobilized on a solid matrix, i.e.2% agarose, it loses over 95% of its inhibitory capacity. The immobilized anti-VIIIR:Ag binds stoichiometrically the antigen and the activity of plasma factor VIII. These results together suggest that factor VIII is composed of 2 different entities, but undissociated under physiological conditions. Immunophysical analyses as a function of pH and temperature of anti-VIII:C and its complex with factor VIII show properties similar to those of homologous antibodies. The antigen determinants of VIII:C (VIII:CAg) are destroyed at low pHs or high temperatures, and VIII:C can no more form a complex with anti-VIII:C. Purified anti-VIII:C is also used in a two-stage assay to detect VIII:CAg or cross-reacting material in some severe haemophiliacs.

Keywords

Antibodies in solid and liquid phase - Neutralization of factor VIII - Immunological stability - Cross-reacting material
 

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