Studies on scale-up parameters of an immunoglobulin separation system using protein A affinity chromatography.
Effects of operational and system parameters on process scale-up of murine immunoglobulin (IgG2a kappa) purification using Protein A affinity chromatography are investigated. Parameters studied are those related to sample application, elution, ligand concentration on support, and column size change. Between sample application velocities of 0.004 and 0.030 cm/s (16-108 cm/h), the product concentration profiles in eluate did not show significant differences. With given system parameters, the retention time and bandwidth of the peak could be predicted by moment theory. The mean equilibrium constant during elution showed a strong effect of pH between 2 and 4. Within the range of protein A concentrations studied, 0.15-1.22 mg/mL of gel, the column capacity shows a linear relationship with the concentration of protein A immobilized. Dimensional scale-up of the column in the radial direction increased the total purification capacity linearly as the cross-sectional area increased without increasing pressure drop; however, the product concentration was diluted. Scale-up of the column dimension in the axial direction enables higher concentrations of product in the eluate, although the retention time increases linearly as the gel height increases.
Kang
K A
KA
Department of Chemical Engineering, University of California, Davis 95616.
Ryu
D D
DD
D000327:Adsorption; D000818:Animals; D002021:Buffers; D002846:Chromatography, Affinity / Q000379:methods*; D005782:Gels; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000302:isolation & purification*; D007700:Kinetics; D008024:Ligands; D051379:Mice; D012685:Sepharose / Q000031:analogs & derivatives; D013205:Staphylococcal Protein A / Q000737:chemistry*
D002021:Buffers; D005782:Gels; D007074:Immunoglobulin G; D008024:Ligands; D013205:Staphylococcal Protein A; D012685:Sepharose; C071734:actigel