A Technique to Purify Monoclonal Antibodies Produced by Chinese Hamster Ovary Cells

Take an FPLC column containing a porous glass resin with immobilized Protein A — an antibody-binding bacterial protein. Equilibrate the column with a neutral pH buffer.

Add cell culture fluid harvested from Chinese hamster ovarian cells, containing monoclonal antibodies and contaminating proteins and DNA.

Protein A binds to the antibody's Fc region, immobilizing them.

Some contaminants non-specifically attach to the resin, while the rest flow through.

In the chromatogram, an increased UV absorbance during sample loading indicates a high protein concentration in the flow through.

Wash with the equilibration buffer to remove unbound contaminants, indicated by a decrease in the absorbance.

Flush with a high-salt buffer, disrupting the non-specific interactions and removing the contaminants, indicated by a small impurity peak.

Add a low-pH elution buffer to disrupt the Protein A-antibody interaction and elute the antibodies, which are collected by tracking the corresponding peak.

Neutralize the pH of the collected antibodies to prevent degradation.