Purification of monoclonal antibodies from clarified cell culture fluid using Protein A capture continuous countercurrent tangential chromatography

• Published in: Journal of biotechnology Vol. 213; pp. 54 - 64
• Main Authors: Dutta, Amit K., Tran, Travis, Napadensky, Boris, Teella, Achyuta, Brookhart, Gary, Ropp, Philip A., Zhang, Ada W., Tustian, Andrew D., Zydney, Andrew L., Shinkazh, Oleg
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 10-11-2015
• Subjects: Animals; Antibodies; Antibodies, Monoclonal - chemistry; Antibodies, Monoclonal - isolation & purification; Binding; Biotechnology; Cell Culture Techniques; CHO Cells; Chromatography; Chromatography - methods; Continuous; Cricetinae; Cricetulus; Monoclonal antibodies; Monoclonal antibody; Productivity; Protein A; Purification; Staphylococcal Protein A - chemistry; Monoclonal antibody; Protein A; Continuous; Chromatography
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2015.02.026

Recent studies using simple model systems have demonstrated that continuous countercurrent tangential chromatography (CCTC) has the potential to overcome many of the limitations of conventional Protein A chromatography using packed columns. The objective of this work was to optimize and implement a CCTC system for monoclonal antibody purification from clarified Chinese Hamster Ovary (CHO) cell culture fluid using a commercial Protein A resin. Several improvements were introduced to the previous CCTC system including the use of retentate pumps to maintain stable resin concentrations in the flowing slurry, the elimination of a slurry holding tank to improve productivity, and the introduction of an “after binder” to the binding step to increase antibody recovery. A kinetic binding model was developed to estimate the required residence times in the multi-stage binding step to optimize yield and productivity. Data were obtained by purifying two commercial antibodies from two different manufactures, one with low titer (∼0.67g/L) and one with high titer (∼6.9g/L), demonstrating the versatility of the CCTC system. Host cell protein removal, antibody yields and purities were similar to those obtained with conventional column chromatography; however, the CCTC system showed much higher productivity. These results clearly demonstrate the capabilities of continuous countercurrent tangential chromatography for the commercial purification of monoclonal antibody products.