Rapid, two-step purification process for the preparation of pyrogen-free murine immunoglobulin G 1 monoclonal antibodies

Abstract

A cost-efficient process was specifically designed for the preparation of gram amounts of highly pure murine immunoglobulin (Ig) G 1 monoclonal antibodies (mAbs). This rapid, simple and scalable purification process employs a unique binding and elution protocol for IgG 1 mAbs on a silica-based, mixed-mode ion-exchange resin followed by conventional anion-exchange chromatography. mAbs are bound to BakerBond ABx medium at pH 5.6 directly from serum-supplemented hybridoma culture supernatants. Contamining proteins and nucleic acids are removed by an intermediate wash at pH 6.5, followed by the specific elution of IgG 1 mabs with 100 m M Tris-HCL (pH 8.5). The mAb eluate is then loaded directly on to QAE-Sepharose Fast Flow medium and eluted with 10 m M sodium phosphate buffer (pH 7.4), containing 150 m M sodium chloride. The resulting IgG 1 mAbs are greater than 98% pure, free from measurable endotoxin, formulated in a physiological buffer and suitable for in vivo applications.