EPITOPE MAPPING OF HUMAN COAGULATION FACTOR VIII WITH IN VITRO SYNTHESIZED FRAGMENTS OF THE PROTEIN
- Resource Type
- Authors
- M Ph Verbeet; A Leyte; R F Evers
- Source
- XIth International Congress on Thrombosis and Haemostasis.
- Subject
- Epitope mapping
Chemistry
Human coagulation factor VIII
Molecular biology
In vitro
- Language
- ISSN
- 2567-689X
We have used recombinant DNA techniques to map the epitopes of the coagulation Factor VIII for several monoclonal antibodies, raised against this protein. For this purpose, we cloned full- length- and partial Factor VIII cDNA sequences in the vector pSP64. Corresponding RNA fragments were synthesized in vitro with SP6 RNA polymerase and translated in a rabbit reticulocyte lysate system. The resulting Factor VII. I polypeptide fragments were immunoprecipitated. We have located the binding sites of a panel of monoclonal anti-Factor VIII antibodies. Two examples are shown in the figure below. The epitopes for anti-Factor VIII CLB-9 and CLB-65 have been confined to areas of 29 (711-740) and 142 (1635-1777) amino acids, respectively. The results of these studies will be useful in determining structure-function relationships of Factor VIII