Protein A chromatography (PAC) is commonly used as an efficient capture step in monoclonal antibody (mAb) separation processes. As PAC media are expensive, it is needed to choose the right column volumn and dimension to process the sample feed within an assigned working time. Usually dynamic binding capacity (DBC) is used for choosing the right PAC. In addition, the elution step is important. For example, if aggregates can be efficiently removed during elution, it can make the following polishing steps easier. For DBC, we have shown a simple dimensionless parameter dp2/(DmRT) is useful for determining the column dimension and the residence time RT (dp:particle diameter, Dm:molecular diffusivity). We have also shown that choosing the right PAC meida in terms of mAb aggregate is possible by using linear pH gradient elution experiments. Usually, aggregates are more strongly retained compared with the monomer. One mAb tested showed different elution behavior, where the aggregates were eluted as both the weakly and strongly retained peaks. In order to remove the two types of aggregates by stepwise elution two protocols were tested. Protocol A consisted of the sample loading, the wash with the equilibration buffer and the low pH elution. The wash stage of protocol B included the wash with 1.0 M arginine. No detectable peaks were observed during the wash stage of protocol A whereas significant peaks were monitored during the arginine wash of protocol B. One of the PAC columns showed a smaller peak during the arginine wash. In addition, both the aggregate removal and the monomer yield were higher with protocol B compared with the other PAC columns. The present method was found to be useful for choosing the right PAC column.

Choosing the right protein A affinity chromatography column can improve the process performance

253rd National Meeting (2017)

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