Epitope mapping of antibodies directed against factor VIII. Recombinant factor VIII fragments corresponding to the factor VIII heavy chain (90K), the A2 domain (A2), and the factor VIII light chain (80K) were expressed in High Five cells and metabolically labeled as described. Binding of anti-factor VIII antibodies to the metabolically labeled factor VIII fragments was assessed by immunoprecipitation and analyzed under reducing conditions on a 10% (wt/vol) SDS-polyacrylamide gel. Lane 1, MoAb CLB-CAg 9 (90K, A2) or CLB-CAg 117 (80K); lane 2, anti-factor VIII antibodies present in the patient's plasma (sample from 1992); and lane 3, control plasma. Immunoprecipitation of the metabolically labeled factor VIII heavy chain gave rise to a nonspecific band of approximately 40 kD. Molecular weight markers are given on the right of the figure.