Purification of Human Coagulation Factors II, IX, and X Using Sulfated Dextran Beads

Joseph P. Miletich, George J. Broze, Philip W. Majerus

Research output: Contribution to journalArticle

59 Scopus citations

Abstract

This chapter describes the production of a highly sulfated derivative of sulfated dextran beads, and its use to isolate human plasma coagulation factors. For production of dextran beads chlorosulfonic acid are added to dry pyridine. The Sephadex G-50 is then added and beads are absorbed on nearly all of the liquid. Further, sodium hydroxide is dissolved in water then rapidly added with mixing to neutralize the acid and beads were collected on a sintered glass funnel and washed. Purification of factors II, IX, and X involve: barium citrate adsorption, ammonium sulfate elution, and dextran sulfate chromatography. Fractions containing the activities of factors II, X, or IX were pooled and concentrated using PM-10 membranes. After 24-hr dialysis at 4°C against 100 volumes of 0.6 M NaC1-0.01 M sodium phosphate (pH 7.0)- 0.001 M benzamidine, the factor pools were filtered and stored in 1-ml portions at –70°C.

Original languageEnglish
Pages (from-to)221-228
Number of pages8
JournalMethods in enzymology
Volume80
Issue numberC
DOIs
StatePublished - Jan 1 1981

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