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Choosing the right protein A affinity chromatography media can remove aggregates efficiently

11:57 EDT 23 Sep 2016 | Wiley Biotechnology Journal

Protein A chromatography (PAC) is commonly used as an efficient capture step in monoclonal antibody (mAb) separation processes. Usually dynamic binding capacity is used for choosing the right PAC. However, if aggregates can be efficiently removed during elution, it can make the following polishing steps easier. In this study a method for choosing the right PAC meida in terms of mAb aggregate removal is proposed. Linear pH gradient elution experiments of two different mAbs on various PAC columns were carried out, where the elution behavior of aggregates as well as the monomer was measured. Aggregates of one mAb were more strongly retained compared with the mAb monomer. Another mAb 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. The first protocol A consisted of the sample loading, the wash with the equilibration buffer and the low pH elution. The wash stage of the second 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. This method was found to be useful for choosing the right PAC column.

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