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Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay
Published on: September 9, 2012
Factor VIII heavy chain polypeptides in plasma and concentrates
G Kemball-Cook1, S A Bevan, T W Barrowcliffe
1Division of Haematology, National Institute for Biological Standards and Control, South Mimms, Potters Bar, Herts.
British Journal of Haematology
|September 1, 1990
Summary
This study analyzed Factor VIII (FVIII) polypeptides using electrophoresis. FVIII in plasma shows larger forms, while concentrates exhibit degradation, varying with manufacturing processes.
Area of Science:
- Biochemistry
- Hematology
- Protein analysis
Background:
- Factor VIII (FVIII) is crucial for blood coagulation.
- Understanding FVIII structure in plasma and concentrates is vital for hemophilia treatment.
- Current analysis methods may not fully capture FVIII heterogeneity.
Purpose of the Study:
- To analyze Factor VIII (FVIII) polypeptide structure in plasma and various FVIII concentrates.
- To compare the integrity of FVIII across different purification and manufacturing processes.
- To identify degradation patterns in FVIII concentrates.
Main Methods:
- Electrophoretic analysis of Factor VIII (FVIII) polypeptides.
- Complexation with radiolabeled anti-FVIII Fab' fragments.
- Visualization of immunocomplexes using SDS-polyacrylamide gel electrophoresis.
- Analysis of plasma, cryoprecipitate, intermediate-purity, highly-purified, and recombinant FVIII concentrates.
Main Results:
- Native FVIII in plasma exhibits heterogeneous, higher molecular weight (Mr) polypeptides (250-300 kDa).
- FVIII concentrates show significant degradation, with reduced high Mr bands and increased lower Mr bands (80-90 kDa, and ~40-50 kDa in some).
- Viral inactivation (heat treatment) and recombinant production methods also resulted in degraded FVIII profiles.
Conclusions:
- Native Factor VIII (FVIII) in plasma is predominantly composed of higher molecular weight forms.
- Concentrate manufacturing processes lead to varying degrees of FVIII degradation.
- The observed degradation impacts the structural integrity of FVIII in therapeutic products.
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