Seeking Innovative Affinity Approaches: A Performance Comparison between Magnetic Nanoparticle Agglomerates and Chromatography Resins for Antibody Recovery

Abstract

Monoclonal antibodies are key molecules in medicine and pharmaceuticals. A potentially crucial drawback for faster advances in research here is their high price due to the extremely expensive antibody purification process, particularly the affinity capture step. Affinity chromatography materials have to demonstrate the high binding capacity and recovery efficiency as well as superior chemical and mechanical stability. Low-cost materials and robust, faster processes would reduce costs and enhance industrial immunoglobulin purification. Therefore, exploring the use of alternative materials is necessary. In this context, we conduct the first comparison of the performance of magnetic nanoparticles with commercially available chromatography resins and magnetic microparticles with regard to immobilizing Protein G ligands and recovering immunoglobulin G (IgG). Simultaneously, we demonstrate the suitability of bare as well as silica-coated and epoxy-functionalized magnetite nanoparticles for this purpose. All materials applied have a similar specific surface area but differ in the nature of their matrix and surface accessibility. The nanoparticles are present as micrometer agglomerates in solution. The highest Protein G density can be observed on the nanoparticles. IgG adsorbs as a multilayer on all materials investigated. However, the recovery of IgG after washing indicates a remaining monolayer, which points to the specificity of the IgG binding to the immobilized Protein G. One important finding is the impact of the ligand-binding stoichiometry (Protein G surface coverage) on IgG recovery, reusability, and the ability to withstand long-term sanitization. Differences in the materials’ performances are attributed to mass transfer limitations and steric hindrance. These results demonstrate that nanoparticles represent a promising material for the economical and efficient immobilization of proteins and the affinity purification of antibodies, promoting innovation in downstream processing.

Magnetic Separation of Antibodies with High Binding Capacity by Site-Directed Immobilization of Protein A-Domains to Bare Iron Oxide Nanoparticles

“…Even if thin and homogeneous coatings are possible, the processing of the particles increases their size compared to bare iron oxide nanoparticles (BION) and decreases their magnetization. 26 To the best of our knowledge, only the work of Iype et al 22 and Padwal et al 20 describes unmodified or nonactivated magnetic nanoparticles as the solid support for the IgG binding ligand. To avoid coating the particles, Iype et al 22 embedded bare magnetic nanoparticles in cross-linked Protein A.…”

Tag-mediated single-step purification and immobilization of recombinant proteins toward protein-engineered advanced materials

“…In the scope of His-tag purification processes, creative approaches are currently being explored by expanding and adapting the IMAC principle to achieve a faster and more efficient purification process without the need for an expensive and laborious column system. One of those innovative systems rely on the use of nanoparticles, mainly magnetic nanoparticles (MNPs) composed of magnetic elements (e.g., iron, nickel or cobalt) and their oxides (e.g., magnetite, maghemite or cobalt ferrite), and a tag attached to the protein of interest that interacts with the MNPs' surface (coated or not) through reversible interactions [39] , [97] , [108] , [109] , [110] , [111] . MNPs encompass several advantages, such as easy manufacture and manipulation, small size, stability and reusability.…”

“…MNPs encompass several advantages, such as easy manufacture and manipulation, small size, stability and reusability. Nanoparticles with or without a magnetic core usually show good biocompatibility (e.g., non-immunogenicity and resistance to plasma protein opsonization) and good biodegradation properties, nevertheless these properties often depend on the synthesis method employed [110] , [111] , [112] . Due to their unique physicochemical properties and versatility, MNPs have potential roles in a number of scientific and technological areas, including in biomedical (e.g., disease diagnosis and therapy), bioengineering (e.g., biosensor and bioelectrode), bioremediation (e.g., water treatment) and biotechnological (e.g., material functionalization and biomolecule immobilization and purification) applications [110] , [112] .…”

Recent trends in sorbents for bioaffinity chromatography

“…A performance comparison for antibody recovery was performed between magnetic nanoparticle agglomerates and chromatography resins functionalized with protein G [139]. A temperature‐responsive affinity chromatography system was constructed for antibody separation using the sorbents with temperature‐responsive dendrimer polyamidoamine‐poly(N‐isopropylacrylamide) as the spacer arms and 4‐mercaptoethylpyridine as the ligand [140].…”