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Research Article Free access | 10.1172/JCI107022
1Hematology Service, Clinical Center, and the Clinical Hematology Branch, National Institute of Arthritis and Metabolic Diseases, National Institutes of Health, Bethesda, Maryland 20014
Find articles by Marchesi, S. in: JCI | PubMed | Google Scholar
1Hematology Service, Clinical Center, and the Clinical Hematology Branch, National Institute of Arthritis and Metabolic Diseases, National Institutes of Health, Bethesda, Maryland 20014
Find articles by Shulman, N. in: JCI | PubMed | Google Scholar
1Hematology Service, Clinical Center, and the Clinical Hematology Branch, National Institute of Arthritis and Metabolic Diseases, National Institutes of Health, Bethesda, Maryland 20014
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Published August 1, 1972 - More info
Factor VIII (antihemophilic globulin) has been prepared from Hyland method IV AHG and cryoprecipitate using limited chymotryptic digestion followed by Sepharose gel filtration. The activity of factor VIII is unaffected by the digestion procedure, while fibrinogen in converted to large noncoagulable fragments.
The purified factor VIII has been found to be a macromolecular glycoprotein with a major subunit of 240,000, as shown by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Carbohydrate analysis of factor VIII gave values of 1% sialic acid, 2.8% hexosamine, and 1-2% hexose (mannose, galactose, and fucose). The lipid content was found to be less than 5% of the protein content, and included no detectable phospholipid. The amino acid content is also reported. Immunoelectrophoretic analysis using rabbit antibody to purified factor VIII produced a single precipitin line.
The chymotrypsin digestion step facilitates the preparation of factor VIII by reducing the viscosity of fibrinogen in the crude starting material, thereby increasing fivefold the quantity of material which can be processed at one time. It also improves markedly the resolution between factor VIII and fibrinogen on gel filtration.
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