Purification of monoclonal antibodies from clarified cell culture fluid using Protein A capture continuous countercurrent tangential chromatography.
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 (∼ 0.67 g/L) and one with high titer (∼ 6.9 g/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.
Chromatan Corporation, 200 Innovation Blvd., Suite 260B, State College, PA 16803, United States; Department of Chemical Engineering, The Pennsylvania State University, University Park, PA 16802, United States.
Amit K
AK
Dutta
Chromatan Corporation, 200 Innovation Blvd., Suite 260B, State College, PA 16803, United States.
Travis
T
Tran
Chromatan Corporation, 200 Innovation Blvd., Suite 260B, State College, PA 16803, United States.
Boris
B
Napadensky
Chromatan Corporation, 200 Innovation Blvd., Suite 260B, State College, PA 16803, United States.
Achyuta
A
Teella
Fujifilm Diosynth Biotechnologies, 101 J. Morris Commons Lane, Morrisville, NC 27560, United States.
Gary
G
Brookhart
Fujifilm Diosynth Biotechnologies, 101 J. Morris Commons Lane, Morrisville, NC 27560, United States.
Philip A
PA
Ropp
Regeneron Pharmaceuticals, 777 Old Saw Mill River Road, Tarrytown, NY 10591, United States.
Ada W
AW
Zhang
Regeneron Pharmaceuticals, 777 Old Saw Mill River Road, Tarrytown, NY 10591, United States.
Andrew D
AD
Tustian
Department of Chemical Engineering, The Pennsylvania State University, University Park, PA 16802, United States.
Andrew L
AL
Zydney
Chromatan Corporation, 200 Innovation Blvd., Suite 260B, State College, PA 16803, United States. Electronic address: oleg.shinkazh@chromatan.com.
Oleg
O
Shinkazh
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000302:isolation & purification*; D016466:CHO Cells; D018929:Cell Culture Techniques; D002845:Chromatography / Q000379:methods; D006224:Cricetinae; D003412:Cricetulus; D013205:Staphylococcal Protein A / Q000737:chemistry*
D000911:Antibodies, Monoclonal; D013205:Staphylococcal Protein A
Chromatography; Continuous; Monoclonal antibody; Protein A