title: Immunopurification of human coagulation factor IX using monoclonal antibodies.
abstract: This study examines the suitability of four recently characterised monoclonal antibodies (MAbs) for the immunoaffinity purification of human coagulation factor IX (FIX) from plasma concentrates. Initial studies using 125I-FIX indicated that appreciable amounts of bound FIX could be eluted from immobilised MAbs with 0.2 M glycine, 50% ethanediol pH 10 (buffer N). Further studies with FIX concentrates showed that buffer N eluted FIX without compromising the activity of the zymogen. Although FIX was eluted from all four MAbs with this buffer, the best yield (82%) was obtained with MAb ESN-3. ESN-3 bound 40 to 60 iu FIX per mg MAb when immobilised on Sepharose 4B. After washing, column elution with buffer N yielded FIX at 100-200 iu/mg. The purity of the product was confirmed by sodium-dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and by Western blotting. The product contained no detectable mouse IgG (less than 3%) and less than 1% FII, X, or protein C.
mesh_terms: D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D004591:Electrophoresis, Polyacrylamide Gel; D005164:Factor IX / Q000302:isolation & purification*; D006801:Humans; D011863:Radioimmunoassay
chemical_list: D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D005164:Factor IX