Chromatographic preparation of a therapeutic highly purified von Willebrand factor concentrate from human cryoprecipitate.
A therapeutic highly purified von Willebrand factor (vWF) concentrate has been prepared from cryoprecipitate by a three-step chromatographic procedure. After solvent/detergent treatment to inactivate viruses, the cryoprecipitate solution was chromatographed on DEAE-fractogel TSK 650 M to separate vWF from most cryoprecipitate proteins, including factor VIII (FVIII) and fibrinogen. A second DEAE-fractogel TSK 650 M was then performed to further purify vWF and to allow concentrating it to over 100 U ristocetin cofactor activity/ml. The last step on immobilized gelatin removed fibronectin and increased the purity of vWF. vWF was recovered with about 18 and 40% yield in antigen and collagen-binding (CB) activity, respectively, from cryoprecipitate. vWF was obtained in an essentially pure state corresponding to a purification factor of over 10,000-fold from plasma. Immunonephelometric and SDS-PAGE analyses of the concentrate did not reveal any detectable cryoprotein contaminants, especially fibrinogen, fibronectin, immunoglobulins and albumin. The content in intermediate- and high-molecular-weight multimers in the concentrate was similar or higher than that of plasma, as the ion-exchanger selectively favored the binding and concentration of the larger multimeric forms while reducing the amount of the smaller forms with abnormal structure and low activity. Other characteristics of the concentrate included a CB activity to antigen ratio of 1.69 and a high capacity (86%) to correct platelet adhesion in a perfusion system. Clinical use of this standardized vWF concentrate has been shown to be efficacious in the treatment of vWF patients.
Burnouf-Radosevich
M
M
Centre Régional de Transfusion Sanguine de Lille, France.
Burnouf
T
T
D000998:Antiviral Agents; D001704:Biopolymers; D001779:Blood Coagulation Factors / Q000032:analysis; D001798:Blood Proteins / Q000032:analysis; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D015925:Cryopreservation; D004591:Electrophoresis, Polyacrylamide Gel; D005169:Factor VIII / Q000302:isolation & purification; D006801:Humans; D015151:Immunoblotting; D010973:Platelet Adhesiveness / Q000502:physiology; D014841:von Willebrand Factor / Q000302:isolation & purification*
D000998:Antiviral Agents; D001704:Biopolymers; D001779:Blood Coagulation Factors; D001798:Blood Proteins; D014841:von Willebrand Factor; D005169:Factor VIII