Sponsored content brought to you by
A long with increases in upstream productivity, next-generation antibody complexity continues to grow. Yet higher product quantities entering downstream processing (DSP) can increase purification complexity without necessarily increasing final product output.
Today’s immunotherapies encompass more sophisticated structures than standard monoclonal antibodies (mAbs). Importantly, these next-generation formats do not always bind to Protein A as expected. For example, they can be pH sensitive or have higher than normal aggregate levels. As a result, the familiar platform solutions may not enable the required specificity or quality.
Without corresponding improvements in chromatography performance, DSP operations can become rate-limiting, creating bottlenecks that offset the manufacturing benefits of achieving higher titers upstream. As protein loads and impurity levels increase, DSP processes must maintain high recovery, consistent product quality, and efficient throughput without sacrificing robustness or scalability.
The purification conundrum
Many, if not most, of the modern antibody formats have the Fc region, and Protein A does work. But, sometimes the antibodies do not bind at all, or low yields result. Additionally, co-elution of product-related impurities that contain the same Protein A binding site can occur. Structural sophistication has mismatched the capabilities of traditional purification methods; DSP, a bottleneck, needs to be re-examined.
The physicochemical properties of these diverse antibody modalities require different purification strategies. For instance, next-generation moieties can have an absent or altered Protein A binding site, be pH sensitive, form dimers or aggregates, have elevated levels of high and low molecular weight species, have product-related impurities that exhibit similar purification behavior as the target molecule, or overexpress light chains and light chain dimers.
Instead of forcing increasingly diverse molecules through a single platform, advanced purification resins can exploit the unique structural characteristics of each molecule.
Utilizing antibody complexity
Process development scientists are increasingly adopting subdomain-specific ligands as a more targeted affinity approach to purify modalities that lack or have altered Fc regions, have other structural enhancements that render them unsuitable for Protein A affinity chromatography, or present product-related impurities that co-elute. Light-chain binding avidity can also help reduce homodimers at the capture step.
Overall, subdomain-specific affinity ligands that focus on specific regions of the antibody beyond the Fc domain offer high selectivity, allowing efficient capture of diverse antibody formats and enhancing both yield and purification performance. These chromatographic ligands can enable robust, scalable purification suitable for cGMP-compliant manufacturing workflows.
Expanding the affinity and polishing toolbox
As manufacturing demands continue to evolve, so do high-performance solutions. Thermo Scientific™ CaptureSelect™ affinity resins help provide highly selective capture for a broad range of antibodies and recombinant proteins, while Thermo Scientific™ POROS™ chromatography resins use convective pore architecture and a rigid backbone to support high flow rates and help process developers improve productivity, streamline purification workflows, and maintain consistent performance.
Based on VHH ligands, CaptureSelect affinity resins are engineered to bind specific regions of human antibodies, such as CH1 and CH3 domains, kappa or lambda light chains, as well as different epitopes than Protein A on the Fc region. The ligands’ small size provides access to binding sites, allowing for highly selective interactions that can increase purity and yield in a single step. Notably, the resins are designed to operate at higher pH than Protein A resins to accommodate sensitive next-generation antibodies.
Post-capture polishing with POROS resins can help provide the final resolution to reduce stubborn impurities from novel mAbs and support the purity and scalability requirements of antibody pipelines.
As titers rise and antibody architectures diversify, the future of purification may be less about finding a single, universal platform and more about flexing the affinity and polishing toolbox to build a process that meets the downstream challenges posed by greater upstream productivity and antibody complexity.
CaptureSelect and POROS resins are for research use only. Not for use in diagnostic procedures.
Tame your misbehavin’ mAbs
thermofisher.com/us/en/home/o/c/bioprocessing-purification-misbehaving-mabs.html
