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Bacterial expression and purification of C1C2 domain of human factor VIII

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With the aim of the production of human factor VIII antigen and its corresponding antibody an epitope coding fragment of the light-chain of hFVIII, fused to a His6-tag, was isolated and over-expressed in Eschehchia coli. The over-expressed hFVIII-epitope containing peptide was confirmed by its reac- tion with a rabbit serum directed against native hFVIII as well as antiHis6-tag antibody. An expression level of 6.5 mg/l (of culture) of the C1C2-related peptide was estimated. The purified product was used to develop antibody in rabbit The immunoblotting experiment confirmed that the rabbit poly- clonal antibodies developed against the purified bacterially expressed hFVIII sub-fragment, recognizes human plasma derived FVIII. Both the produced hFVIII-related antigen and its corresponding antibody are useful in experiments using for detection and purification of hFVIII as well as the clinical diagnosis of hFVIII related disorders.

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