A MONOCLONAL-ANTIBODY TO VIII-C PRODUCED BY A MOUSE HYBRIDOMA

  • 1 January 1981
    • journal article
    • research article
    • Vol. 58  (5) , 1000-1006
Abstract
Spleen cells of a BALB/c mouse immunized with human factor VIII (FVIII) procoagulant activity (VIII:C) (isolated by affinity chromatography) were fused with mouse myeloma cells (P3 .times. 63 Ag8). After the fusion, 12/32 wells produced an inhibitor to VIII:C. Cells from 1 well (1B3) were subcloned 4 times to isolate the hybridoma that produces the anti-VIII:C antibody. Injection of hybridoma cells in pristane pretreated BALB/c mice results in anti-VIII:C titers of 5000-10,000 Bethesda U/ml. Analysis of the produced Ig demonstrated H chains of IgG1 (produced by the myeloma cell line) and IgG2b subclass. The 1B3 antibody neutralizes VIII:C in the low MW FVIII fraction, crysosupernatant, cryoprecipitate and normal plasma. Binding of the IgG to FVIII results in a delay in its activation and not in an inhibition of its cofactor activity. The antibody removes VIII:C from pooled normal plasma when coupled to Sepharose; when coupled to plastic tubes, it binds VIIICAG [VIII:C related antigen] from isolated VIII:C, purified FVIII and pooled normal plasma; it does not bind VIIIR:AG [FVIII-related antigen], fibrogen or serum VIIICAG. The 1B3 antibody can be used successfully in an immunoradiometric assay for VIIICAG.