Antibody Aggregate Removal by Multimodal Chromatography

Abstract

The growing demand for therapeutic monoclonal antibodies (mAbs) has heightened the need for efficient and scalable purification strategies. A major challenge in downstream processing is the removal of antibody aggregates, which can compromise drug safety, efficacy, and regulatory compliance. This review explores the use of multimodal chromatography for aggregate separation, providing an in-depth analysis of commercially available resins and emerging adsorbent prototypes. It also examines the mechanisms of aggregate formation during bioprocessing. A comparative evaluation of conventional single-mode chromatography techniques-affinity, ion exchange, and hydrophobic interaction-is presented alongside multimodal chromatography, which integrates ion-exchange, hydrophobic, and other non-covalent interactions for enhanced aggregate clearance and process flexibility. The review primarily assesses commercial multimodal resins in terms of aggregate removal efficiency, binding capacity, and scalability. Additionally, advancements in prototype resins and multimodal membranes are discussed. Finally, the advantages, limitations, and future directions of multimodal chromatography in mAb aggregate removal are outlined. As purification demands continue to evolve, multimodal chromatography is poised to play an increasingly critical role in achieving the high purity standards required for therapeutic antibodies.

Keywords: aggregate formation; antibody aggregates; hydrophobic interaction chromatography; ion-exchange chromatography; mAb purification; multimodal chromatography.

Figure 1

Schematic representation of the antibody aggregation process. The red arrows represent the non-native aggregation, while the dark blue arrows denote the native aggregation. The bidirectional arrows show the reversible steps, and the mono-directional arrows account for the irreversible process [11].

Figure 2

Hofmeister series of anions and cations and their influence on protein solution properties. Upward (↑) and downward (↓) arrows indicate an increase or decrease, respectively, of specific protein property [83].

Figure 3

The structure of hydroxyapatite.

Figure 4

Chemical structures of the Capto Adhere (A), Capto Core 700 (B), PPA HyperCel (C), HEA HyperCel (D), Nuvia aPrime 4A (E) and MEP HyperCel (F) ligands.

Figure 5

Chemical structures of the Capto MMC (A), Poros Caprylate (B), Toyopearl MX-Trp-650M (C), Eshmuno HCX (D), Eshmuno CMX (E), and Nuvia cPrime (F) ligands.