Purification of six human vitamin K-dependent proteins in a single chromatographic step using immunoaffinity columns

Preparative Biochemistry
R JennyK G Mann

Abstract

The major human vitamin K-dependent proteins were purified from plasma using immunoadsorbents made with antibodies specific for each protein. Monoclonal antibodies to Factor VII, Factor IX, Factor X, Protein C, and Protein S were prepared from mice immunized with isolated vitamin K-dependent antigens. Purified monoclonal antibodies and a purified burro polyclonal anti-prothrombin immunoglobulin were individually coupled to Sepharose and used in a tandem series of columns to purify each of the vitamin K-dependent proteins from eluates of barium citrate precipitates of plasma. The proteins were eluted from the columns by sodium thiocyanate and retained functional activity following dialysis. Prothrombin, Factor VII, Factor IX, Factor X and Protein C were essentially homogeneous as judged by NaDodSO4-PAGE; Protein S was isolated as a Protein S-C4b binding protein complex. These results indicate the utility of monoclonal antibody immunoadsorbents for purifying the human vitamin K-dependent proteins and represent a considerable simplification over other purification schemes.

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Citations

Mar 1, 1996·Thrombosis Research·W T SumnerM Hoffman
Jun 8, 2001·Journal of Chromatography. B, Biomedical Sciences and Applications·H Husi, M D Walkinshaw
Feb 17, 2000·Journal of Chromatography. B, Biomedical Sciences and Applications·H Husi, M D Walkinshaw
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Jun 18, 2010·Arteriosclerosis, Thrombosis, and Vascular Biology·Kenneth G Mann
May 25, 2010·Journal of Thrombosis and Haemostasis : JTH·T OrfeoK G Mann
Jan 1, 1990·The American Journal of Medicine·W T Branch, A Suchman
Dec 24, 2013·Biochimica Et Biophysica Acta·Beth A BouchardSaulius Butenas
Nov 30, 2007·Academic Medicine : Journal of the Association of American Medical Colleges·Larry A GreenPerry A Pugno
Feb 25, 2011·Academic Medicine : Journal of the Association of American Medical Colleges·Nicholas Kontos
Apr 18, 1994·Annals of the New York Academy of Sciences·K G Mann

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