Monoclonal antibodies to human procoagulant factor VIII.
VIII:C was purified from intermediate-purity concentrate by adsorption on polyelectrolyte E5 and affinity chromatography on Sepharose/anti-VIIIR:Ag. The highly purified VIII:C preparation (sp. act. 1598 U/mg) was used to immunize Balb-C mice. Spleen cells from a mouse with a serum antibody titer of 963 U/ml were fused with P3 NSI mouse myeloma cells. Hybrid clones were screened by a coagulation inhibition assay and by a four-layer antibody adsorption procedure. Nine monoclonal antibodies specific to VIII:C were produced. Five of these antibodies have been cloned and grown in mouse ascitic fluid. Antibody titers from ascitic fluid ranged from 35 to 82,000 BU/ml. The antibodies, when radiolabeled, form a high-molecular-weight complex with antigens present in normal plasma and factor VIII concentrate, but not when incubated with CRM-negative hemophilic plasma. A two-site assay using a combination of monoclonal antibodies is able to detect VIII:CAg in normal plasma and in factor VIII concentrate. Sensitive two-site immunoradiometric assays using monoclonal antibodies as the solid phase have been set up.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D000941:Antigens / Q000276:immunology* / Q000302:isolation & purification; D003429:Cross Reactions; D005169:Factor VIII / Q000276:immunology* / Q000302:isolation & purification; D006467:Hemophilia A / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007074:Immunoglobulin G / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D011863:Radioimmunoassay / Q000379:methods
D000911:Antibodies, Monoclonal; D000941:Antigens; D007074:Immunoglobulin G; D005169:Factor VIII