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

Abstract

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.

Enhancing Protein A performance in mAb processing: A method to reduce and rapidly evaluate host cell DNA levels during primary clarification

“…Other methods to overcome the challenges chromatin/DNA presents to Protein A chromatography involve modifying the operating conditions for Protein A chromatography itself. One strategy used chromatofocusing using a pH gradient through the Protein A column to reduce HCP coelution with the target mAb . An alternative approach made use of two distinct flow rates, high and low, to influence residence time and minimize nonspecific binding and HCP hitchhiking as well as to improve overall column utilization .…”

“…One strategy used chromatofocusing using a pH gradient through the Protein A column to reduce HCP coelution with the target mAb. 18 An alternative approach made use of two distinct flow rates, high and low, to influence residence time and minimize nonspecific binding and HCP hitchhiking as well as to improve overall column utilization. 19 Changes to the column washing strategy have also been considered to improve HCP and other contaminant clearance across Protein A chromatography.…”