TY - JOUR
T1 - Purification of Human Coagulation Factors II, IX, and X Using Sulfated Dextran Beads
AU - Miletich, Joseph P.
AU - Broze, George J.
AU - Majerus, Philip W.
N1 - Funding Information:
This research was supported by Grants HLBI 14147 (Specialized Center in Thrombosis), HL 07088, and HL 16634 from The National Institutes of Health, and in part by NIH Research Service Award GM 07200, Medical Scientist, from the National Institute of General Medical Sciences.
PY - 1981/1/1
Y1 - 1981/1/1
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=0000435625&partnerID=8YFLogxK
U2 - 10.1016/S0076-6879(81)80020-1
DO - 10.1016/S0076-6879(81)80020-1
M3 - Article
AN - SCOPUS:0000435625
SN - 0076-6879
VL - 80
SP - 221
EP - 228
JO - Methods in enzymology
JF - Methods in enzymology
IS - C
ER -