Bispecific Antibody Purification’s Complexity Problem and How to Fix It

Bispecific antibodies, which can target multiple disease mechanisms at once, are transforming the biologics field. However, their complex structures create several challenges during purification, making them more difficult to handle than traditional monoclonal antibodies. Unlike monoclonals, bispecifics often generate a range of product-related variants such as mispaired chains, half antibodies, homodimers, and aggregates. These variants can be hard to separate due to their unique physicochemical properties.

To overcome these issues, analytical insights combined with adaptable process design are crucial. Thermo Fisher Scientific provided several case studies demonstrating how proper analytics can inform the selection and optimization of purification steps.

Firstly, for bispecifics with different light chain types, resins offering high binding capacity for kappa and lambda light chains showed promising results. Further optimization involved using pH-gradient elution and elution modifiers like magnesium chloride to enhance separation and purity, although this sometimes reduced yield.

Secondly, to tackle high levels of aggregates at the capture stage, using a Fc-targeting resin that binds a different epitope than protein A provided better results. This approach significantly improved the purity of the monomer fraction and reduced aggregate levels.

Lastly, leveraging the selectivity of a specific CH1 domain allowed for clearer separation in complex mixtures, leading to high monomer purity and reduced aggregate content. Overall, the key is to "listen to what the analytics are telling us" and make informed decisions about the purification strategies, as the diversity of bispecific structures demands customized approaches.