Separation of human vitamin K-dependent coagulation proteins using hydrophobic interaction chromatography

Husi, H. and Walkinshaw, M. D. (1999) Separation of human vitamin K-dependent coagulation proteins using hydrophobic interaction chromatography. Journal of Chromatography B: Biomedical Sciences and Applications, 736(1-2), pp. 77-88. (doi: 10.1016/S0378-4347(99)00437-5) (PMID:10676986)

Abstract

A rapid and simple method was developed to separate human vitamin K-dependent plasma proteins from each other, yielding virtually homogeneous pools. The purification technique is based on the single use of hydrophobic interaction chromatography, starting from prothrombin concentrate (PC or DEFIX, also termed factor IX concentrate) as initial material. Phenyl-sepharose HP demonstrated optimal separation by comparing several hydrophobic resins as well as resins used in standard procedures like immobilised heparin and Cibacron blue. Under ideal conditions, factor X could be separated in a single step as well as prothrombin. Factor IX co-eluted with other minor proteins. Focus was given only on these three proteins due to their relative abundance. Complete separation of all proteins present in the starting material was achieved by MonoQ anion-exchange chromatography following the phenyl-sepharose run. The resulting purified material could be demonstrated to be of equal or higher purity than using described methods. This strategy employing hydrophobic interaction chromatography for blood macromolecules could be of immense value for purifying the human vitamin K-dependent proteins and represents a considerable simplification over other purification schemes. It not only involves minimal sample handling but also can be readily up-scaled and is a cost-efficient alternative.

• Item Type: Articles (Other)
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Husi, Dr Holger
• Authors: Husi, H., and Walkinshaw, M. D.
• College/School: College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health
• Journal Name: Journal of Chromatography B: Biomedical Sciences and Applications
• Publisher: Elsevier
• ISSN: 1387-2273
• Published Online: 09 December 1999