Low pH treatment is a commonly known factor for the aggregation of antibodies and Fc-Fusion proteins. During downstream processing of these proteins, low pH conditions are used as an effective way for virus inactivation at pH 3.0-3.5, as well as for elution from a previous Protein A chromatography step. Binding to Protein A material is induced through multiple interactions between the Fc-part of the antibody with the immobilized Protein A domains and is sensitive for conformational changes.<br/>The objective of this work was to use low pH treatment to study its effect on aggregation and structural changes of monoclonal antibodies. Therefore, a Protein A purified antibody solution was acidified (pH 2.8 to 3.5) and incubated for a period of time. Afterward, the solution was neutralized and reloaded onto a Protein A chromatography column. This resulted in only a partial binding of the sample; furthermore, when analyzing the resulting elution peak by size exclusion chromatography, a double-peak elution behavior at the monomer size was observed. Both phenomena are independent of each other but occurred during the same experiment. Additionally, these effects could not be nullified through the addition of excipients to the buffer system.<br/>The results show that multiple low pH treatments can significantly change the structure and binding behavior of monoclonal antibodies and should, therefore, be kept at a minimum during purification.

Influence of low pH treatment on the elution behavior of monoclonal antibodies in protein A chromatography

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.