Choosing the right protein A affinity chromatography media can remove aggregates efficiently. Issue 1 (10th October 2016)

• Record Type: Journal Article
• Authors: Yada, Tomokazu; Nonaka, Koichi; Yabuta, Masayuki; Yoshimoto, Noriko; Yamamoto, Shuichi
• Is Part Of: Biotechnology journal. Volume 12:Issue 1(2017)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2016-10-10
• Subjects: Arginine -- Monoclonal antibody -- pH gradient elution -- Protein A chromatography -- Protein aggregates; Biotechnology -- Periodicals; 660.605
• DOI: 10.1002/biot.201600427
• Languages: English
• ISSNs: 1860-6768

Abstract

Abstract: 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 media in terms of mAb aggregate removal is proposed. Linear pH gradient elution experiments of two different mAbs on various PAC columns are carried out, where the elution behavior of aggregates as well as the monomer is measured. Aggregates of one mAb are more strongly retained compared with the mAb monomer. Another mAb showed different elution behavior, where the aggregates are eluted as both the weakly and strongly retained peaks. In order to remove the two types of aggregates by stepwise elution two protocols are 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 are observed during the wash stage of protocol A whereas significant peaks are monitored during the arginine wash of protocol B. One of the PAC columns showed a smaller peak during the arginine wash. In addition, both aggregate removal and monomer yield are higher with protocol B compared with the other PAC columns. This method is found to be useful for choosing the right PAC column. Abstract : Protein A chromatography is commonly used as an efficient capture step in monoclonal antibody separation. By choosing the right protein A chromatography media, different types of aggregates (type 1: normal mAb‐stronger binding and type 2: hybrid and truncated mAb‐weaker binding) can be efficiently removed during the wash and the elution periods. This study proposes a method to select the best media, which can make the following polishing chromatography steps easier.