Monoclonal antibodies to factor VIII: their application in immunoblotting for the visualization of factor VIII in therapeutic concentrates and plasma

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Abstract
Two monoclonal antibodies (McAb) termed 12A4 and 19C1 have been raised against human factor VIII. In immunoassays 12A4 bound to factor VIII antigen (VIII:Ag) in plasma but not serum whilst 19C1 bound to VIII:Ag in both plasma and serum. Both McAb were shown by immunoblotting to react with the carboxy (C) terminal polypeptide of factor VIII which appeared as a doublet with a molecular weight (Mr) of 77,000/75,000. The C terminal factor VIII polypeptide was detectable by immunoblotting in each of 12 therapeutic factor VIII concentrates, from six different manufactures, although its level was variable. Factor VIII was visualized in plasma by immunoblotting following its immunoadsorption and elution from agarose-bound monoclonal antibodies. No Mr 77,000/75,000 bands were detectable in plasma obtained from 13 unrelated CRM- haemophiliacs whilst 11 CRM+ haemophilic plasmas from seven kindred were shown to have a C terminal factor VIII polypeptide of normal molecular size.