Use and optimization of a dual-flowrate loading strategy to maximize throughput in protein-a affinity chromatography.
The effect of an alternate strategy employing two different flowrates during loading was explored as a means of increasing system productivity in Protein-A chromatography. The effect of such a loading strategy was evaluated using a chromatographic model that was able to accurately predict experimental breakthrough curves for this Protein-A system. A gradient-based optimization routine is carried out to establish the optimal loading conditions (initial and final flowrates and switching time). The two-step loading strategy (using a higher flowrate during the initial stages followed by a lower flowrate) was evaluated for an Fc-fusion protein and was found to result in significant improvements in process throughput. In an extension of this optimization routine, dynamic loading capacity and productivity were simultaneously optimized using a weighted objective function, and this result was compared to that obtained with the single flowrate. Again, the dual-flowrate strategy was found to be superior.
Ghose
Sanchayita
S
Purification Process Development, Amgen Inc., Seattle, Washington, USA.
Nagrath
Deepak
D
Hubbard
Brian
B
Brooks
Clayton
C
Cramer
Steven M
SM
D000327:Adsorption; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D000936:Antigen-Antibody Complex / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D016466:CHO Cells; D002846:Chromatography, Affinity / Q000379:methods*; D045424:Complex Mixtures / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D003198:Computer Simulation; D006224:Cricetinae; D003412:Cricetulus; D003470:Culture Media / Q000737:chemistry / Q000302:isolation & purification; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D044085:Microfluidics / Q000379:methods*; D008956:Models, Chemical; D011786:Quality Control; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D013205:Staphylococcal Protein A / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D045424:Complex Mixtures; D003470:Culture Media; D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A