Use and Optimization of a Dual-Flowrate Loading Strategy To Maximize Throughput in Protein-A Affinity Chromatography

Abstract

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.

Maximizing binding capacity for protein A chromatography

“…This work herein demonstrates a practical way of achieving very high DBC (>65 g/L) on Protein A to address the demands of high titer processes. Previous work in the literature had introduced the concept of dual flow loading strategy to increase both productivity and capacity of protein A resins . This work extends the possibility of applying this loading strategy on a new generation protein A media (MabSelect SuRe LX) in an effort to maximize binding capacity on Protein A.…”

“…A dual flowrate loading strategy has been shown in the past to significantly improve productivity in Protein A chromatography . In principle, a faster flowrate (or a lower residence time) was used in the initial stages of loading when all the binding sites are available.…”

A comparison of Protein A chromatographic stationary phases: Performance characteristics for monoclonal antibody purification

“…The ratio of dynamic binding capacity to operation time is important, as this governs the productivity of the process . A dual‐flow‐rate strategy with rapid flow rates initially followed by slower flow was demonstrated to increase throughput on Protein A resins by enabling more complete binding to the stationary phase . It is also important to recognize that these priorities can shift as drug development progresses.…”

Future of antibody purification

“…Within the limitations of existing binding capacity, a strategy to achieve a higher throughput was proposed by Cramer and co-workers [29] using a dual flow rate approach. In the early phase of loading, the flow rate is kept high, since all of the binding sites are readily available for IgG binding.…”

“…Since binding capacity declines at higher flow rates, one could load the column at a higher flow rate initially and then reduce it as maximum capacity is reached [29,96].…”