Evaluation of two-different affinity chromatographic columns based on Protein A for the purification of an IgG2a monoclonal antibody from ascites

Abstract

Purification of monoclonal antibodies (MAbs) is today a routine procedure in laboratories destined for obtaining these molecules. Several approaches, based mainly on differential solubility and chromatographic methods, are frequently used for reaching the purity required of these products. For IgG MAbs, affinity chromatography using immobilized protein A or G, is frequently the best option to perform a purification protocol in a single step. In our laboratory, the purification of MAbs like 7E1F7 (IgG2a) against capsular polysaccharide from Neisseria meningitidis serogroup A, are usually done by affinity chromatography on Protein G. The objective of this work was to evaluate the feasibility to use two-different affinity chromatographic columns based on Protein A (HiTrap from GE and CIM-A from BIA Separations), for the purification of this MAb. The same purification protocol was followed with both columns. Percentage of recovery, purity and immunoreactivity of the purified MAb were calculated and used to compare the performance of both columns. Similar chromatographic profiles were obtained from both columns. However, for CIM-A column, protein concentration and total recovery were significantly higher (p<0.0001). Purity levels of the 7E1F7 MAb over 95% was obtained with both columns without any change on its immunoreactivity. The use of Protein A coupled to monolith supports, like from BIA Separations Company, offers an attractive possibility to purify MAbs from ascites with high recovery, purity levels and fully preserved reactivity. This paper constitutes the first report on the use of monolithic columns in the purification of monoclonal antibodies from ascites.