Protein A affinity chromatography for capture of monoclonal antibodies results in a high purity and yield over one step. In order to utilize the Protein A resin for numerous cycles there is a need to efficiently clean and sanitize it using harsh chemical conditions. This can be a challenge depending on the sensitive nature of the protein based ligand. Development of alkali stabilized Protein A ligands tolerating up to 0.5 M NaOH has improved the ability to clean and sanitize Protein A resins over the last decade. These NaOH concentrations may be sufficient for removal of fouled proteins and reduction of vegetative bacteria, while spore forming bacteria is still a challenge. Thus, Protein A ligand development is underway towards ligands that tolerate even sporicidal concentrations of NaOH. In addition, more efficient sanitization agents, based on oxidizing chemicals, have shown promising results for affinity resins sensitive to high concentrations of NaOH.<br/> <br/>The aim of this work was to study molecular modifications of existing and pipeline Protein A affinity ligands after exposure to both common and novel cleaning and sanitization agents. Amino acid site specific modifications were elucidated by high resolution mass spectrometry and the effect on kinetics and binding affinity was monitored by Surface Plasmon Resonance. Finally, the functionality and lifetime of the Protein A resins after regular exposure to cleaning and sanitization agents were evaluated by confocal microscopy and conventional chromatographic methods. <br/>

Setting new standards for cleaning and sanitization of Protein A chromatography resins

253rd National Meeting (2017)

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