EFFICIENT PURIFICATION OF MOUSE ANTI-FGF RECEPTOR IGM MONOCLONAL-ANTIBODY BY MAGNETIC BEADS

Affinity chromatography has been widely used for the purification of m onoclonal antibodies (MAb), Traditionally, activated agarose beads con jugated with specific antisera have been used as a solid support in ch romatographic protein purification. Magnetic beads conjugated with var ious antibodies have recently become an alternative method for the iso lation of diverse proteins, nucleic acids, and cell types, In this stu dy, murine anti-fibroblast growth factor receptor 1 (FGFR1) immunoglob ulin M (IgM) was isolated from protein solutions to compare immunoaffi nity column chromatography and magnetic bead IgM purification methods, Using immobilized rat anti-mouse IgM MAb, an UltraLink l-ethyl-3-(3-d imethylaminopropyl)carboiimide (EDC)/diaminodipropylamine (DADPA) immu noaffinity column and polystyrene-coated magnetic beads were used for the purification of mouse IgM from bovine serum albumin/phosphate-buff ered saline (BSA/PBS) as well as from crude ascites, Protein quantitat ion and percent IgM yield were determined by reducing SDS-PAGE electro phoresis followed by silver staining, then IgM and protein contaminant s were quantitated using densitometry analysis. IgM anti-FGFR1 binding specificity and immunologic activity were determined by enzyme-linked immunosorbant assay (ELISA), This study demonstrates that magnetic be ad isolation of IgM from ascites is more effective than traditional af finity chromatography purification as determined by greater IgM yield, purity, and immunologic activity.