n-Layer BET adsorption isotherm modeling for multimeric Protein A ligand and its lifetime determination

[Display omitted] •n-layer BET isotherm model explains the binding characteristics of a rProtein A ligand.•Binding capacity decreases with exposure to caustic, except at time zero for new resin.•Adding salt to sanitization regime improves the binding capacity for a limited time.•Static and dynamic adsorption confirms the improved binding with caustic for new resin.•Fundamental representation of a multimeric Protein A ligand is the novelty of this paper. Langmuir and other single-layer adsorption isotherms show the binding behavior of natural Protein A ligands immobilized on a column. However, no models have been shown in literature to explain the adsorption phenomena on the recombinant high binding capacity Protein A resins. This study has characterized the Protein A binding domain distribution across the ligand with multi-layer adsorption isotherms for a recombinant Protein A resin. The adsorption data was analyzed using the Langmuir, Freundlich, Brunauer–Emmett–Teller (BET) and various other mathematical equations. The best fit of experimental data was obtained with n-layer BET model wherein the isotherms of Protein A exhibited Type IV behavior according to BET classification. Furthermore, the binding capacity was studied throughout the shelf life using the multi-layer adsorption isotherm model. Antibody adsorption isotherms of Protein A resin were obtained at preset duration of caustic incubation. The experiments were carried out for two conditions of sanitization agent, namely, caustic and caustic with salt. Static and dynamic isotherm analysis showed that a new resin had a lower binding capacity and the initial sanitization improved the binding capacity, probably by making the binding domains more accessible. The binding capacity at equilibrium, dynamic breakthrough and batch were also evaluated and reported in this paper. The study modeled the multimeric Protein A ligand and established the requirement of optimization for cleaning regime. This study provides a fundamental understanding of the binding patterns in the recombinant Protein A ligands through a working mathematical equation and improves the current knowledge of Protein A resin lifetimes.