Precipitation of process-derived impurities prior to capture chromatography in antibody purification offers a simple, robust,
and economical method to efficiently remove Chinese hamster ovary (CHO) host cell proteins and DNA. By optimizing the major
process parameters—pH, caprylic acid concentration, and mixing time—and understanding their interdependency, one can develop
a cost-effective process step. When precipitation is applied directly in CHO cell culture, it combines the clarification and
precipitation unit operations. The direct precipitation of contaminants results in seamless transition from upstream to downstream
purification processes, particularly in high cell density and high titer cell culture. As a result, demand on the purification
process is significantly lowered, and a simple two-step ion exchange process is sufficient to achieve therapeutic purity.
Cation exchange (CEX) capture chromatography in non-affinity processes generally requires a feed conditioning step to lower
the pH and conductivity in order to attain high binding capacity (~100 mg/mL). Typically, this is achieved by including a
concentration and diafiltration step for primary recovery or, less preferably, through dilution and pH titration of the clarified
cell culture bulk, resulting in large processing volumes.1
Sartorius Stedim Biotech GmbH
Primary recovery TFF offers several advantages, including batch volume reduction, partial purification of process-derived
DNA, and a cleaner feed stream for better column lifetime and performance.1 On the other hand, it is a lengthy unit operation, with high costs in terms of buffer consumption and TFF cassettes. Diafiltration
also adds further constraints in the case of high titers by requiring even higher buffer and tank volumes at production scale.
For those reasons, alternative technologies such as precipitation have been explored, particularly to achieve higher throughput
for cell culture processes with high titers. Precipitation of the protein of interest has proven successful in food, blood,
and enzyme manufacturing. For example, antibodies have been precipitated successfully at large scale by adding polymers of
ethylene glycol (PEG)2 or salts3 and pH titration,4 and research-scale precipitation of contaminants by charged polymers,5 cationic detergents, or short-chain fatty acids6,7 has shown promising application at the cell culture clarified bulk (CB) stage.
In this study, we extend the application of precipitation that combines pH control and caprylic acid (a short-chain fatty
acid) to remove process-derived impurities directly from CHO cell culture in the production of human monoclonal antibodies