A Novel Ligand For The Chromatographic Purification Of Antibodies

The growing role of antibodies in biopharmaceutical research and development is widely acknowledged. The high dosage of therapeutic antibodies and the parenteral use of diagnostic antibodies require that products meet stringent requirements for purity, safety and potency. A novel chromatographic ligand was screened for improvement of the purity of antibodies. The screen principle was based on the characteristic of hydrophobic charge induction chromatography and antibodies'purification. The screened ligand was coupled to the matrix activated with allyl bromide.The mechanism of adsorption was investigated withγglobulin and bovine serum albumin (BSA) as the model proteins. The effects of liquid-phase ionic strength, pH values and additives on the adsorption behaviors of the model proteins were studied in detail. It was foundγglobulin was salt-independent adsorption, but BSA was sensitive to ionic strength. The results indicated the hydrophobic interaction might play the most important role in the adsorption betweenγglobulin and the ligand. While, the electrostatic interactions might be the most important contributor to the BSA adsorption. At pH5, both the ligand andγglobulin carrying a net positive charge, desorption occurred on basis of electrostatic charge repulsion. And because of the difference of the isoelectric point (pI), BSA was still binded to the matrices. The novel-ligand matrices were introduced to separate antibodies from protein mixture and bovine serum. After optimizing the separation conditions, it was found the novel ligand could adsorb the antibodies selectively and effectively, without the interference of albumin. It got the high-purity immunoglobulins (purity >90%) from protein mixture. The purity of immunoglobulin G from the bovine serum was 80% with the purification factor 37.2. In addition, the antibodies adsorbed to the matrices could be recovered efficiently in the milder elution conditions.