Thermodynamics of the adsorption of monoclonal antibodies in phenylboronate chromatography: Affinity versus multimodal interactions

Antibody Binding Heterogeneity of Protein A Resins

“…By reproducing lab‐scale conditions of a regular chromatography system, FMC is a powerful tool to characterize a chromatographic process with respect to the associated thermodynamics. We have already successfully investigated the energy profile in the purification of other biomolecules, namely lysozyme, plasmid DNA, and mAbs, and concluded that biomolecule purification in liquid chromatography is a complex process that should be carefully interpreted. The adsorption of a biomolecule to a ligand in a chromatographic process has different characteristics from a batch process both in terms of mass transfer properties and associated energy.…”

“…FMC, on the other hand, is able, to some extent, to dissect the subprocesses involved in the interaction between the biomolecule and the resin and as a consequence to discriminate between different energy sources. The technique has been used, with good results, to study the adsorption thermodynamics of biomolecules on ion exchange, hydrophobic interaction, and mixed‐mode chromatography . FMC simulates lab‐scale chromatography and implies that the system is in equilibrium.…”

Mobile‐Phase Modulators as Salt Tolerance Enhancers in Phenylboronate Chromatography: Thermodynamic Evaluation of the Mechanisms Underlying the Adsorption of Monoclonal Antibodies

“…An anti‐IL8 mAb (from the immunoglobulin G1 [IgG1] family of immunoglobulins) was obtained by in‐house production using CHO DP‐12 cells grown under serum‐free culture conditions in single or multi‐T‐175 flasks (BD Falcon, Franklin Lakes, NJ, USA), as previously described . The anti‐IL8 mAb presents a pI around 9.3 and its concentration in the serum‐free cell culture supernatants obtained varied between 75 mg L −1 and 100 mg L −1 .…”

“…Nevertheless, a deeper understanding of the phenomena involved in mAbs adsorption combined with the economic advantages of employing the PBA ligand, could change the paradigm in mAbs downstream processing, pushing the industry to adopt such chromatographic process either for mAb capture and/or polishing steps or even as a prechromatographic step for a longer lifetime of the costly protein A resin. Isothermal titration calorimetry (ITC), surface plasma resonance, confocal laser scanning microscopy, attenuated total reflectance–Fourier transform infrared spectroscopy, nuclear magnetic resonance, small‐angle X‐ray scattering, and flow microcalorimetry (FMC) are some of the techniques that have been employed to study the mechanisms behind the adsorption of biomolecules. However, from the above‐mentioned techniques, only few can operate in situ, i.e., in the chromatographic column, and consequently, account directly for the dynamics of the chromatographic process.…”

“…One of these techniques is FMC, which is able to simulate a packed‐bed chromatographic system at a microscale. The heat signal profile (magnitude and chronology of the thermal events) obtained can provide an overview of the driving forces of the adsorption of biomolecules, from small proteins to larger molecules such as mAbs and plasmid DNA …”