Immunoglobulin G elution in protein A chromatography employing the method of chromatofocusing for reducing the co‐elution of impurities

Purification processes for monoclonal Immunoglobulin G (IgG) typically employ protein A chromatography as a capture step to remove most of the impurities. One major concern of the post‐protein A chromatography processes is the co‐elution of some of the host cell proteins (HCPs) with IgG in the capture step. In this work, a novel method for IgG elution in protein A chromatography that reduces the co‐elution of HCPs is presented where a two‐step pH gradient is self‐formed inside a protein A chromatography column. The complexities involved in using an internally produced pH gradient in a protein A chromatography column employing adsorbed buffering species are discussed though equation‐based modeling. Under the conditions employed, ELISA assays show a 60% reduction in the HCPs co‐eluting with the IgG fraction when using the method as compared to conventional protein A elution without affecting the IgG yield. Evidence is also obtained which indicates that the amount of leached protein A present in free solution in the purified product is reduced by the new method. Biotechnol. Bioeng. 2017;114: 154–162. © 2016 Wiley Periodicals, Inc.