This talk presents a case study in the practical application of automation and mechanistic chromatography modeling in early-stage (FIH) purification process development. High-throughput screening and modeling are used in conjuction to support PD and subsequent tech transfer of a cation-exchange polishing step for an mAb with high aggregate burden. Importantly, the model is shown to be useful in adapting operating conditions to changing feed impurity levels due to cell-line improvements, thereby improving process outcomes at scale. Overall, the proposed workflow allowed for timeline acceleration, process improvement, and de-risked tech transfer, enabling the toxicology study to support IND.

The bioprocessing industry is moving toward Bioprocessing 4.0, using digital tools such as mechanistic chromatography models to increase process understanding and efficiency. We developed and calibrated Protein A capture and cation exchange polishing models using linear gradient and step elution experiments with varied load challenges. These models were then applied in a digital workflow to determine proven acceptable ranges and rank process parameters by criticality. Comparison with traditional, experiment-heavy process characterization (PC) demonstrated that mechanistic models can effectively and efficiently guide PC studies, enabling well-defined operating ranges and accelerated, more informed PC campaigns.

Adequate mixing and buffer exchange during ultrafiltration/diafiltration (UF/DF) are critical for robust downstream bioprocess performance, particularly when mechanical agitation is constrained by equipment or operating limits. In this study, computational fluid dynamics (CFD) modeling was applied to evaluate hydrodynamics, mixing efficiency, and buffer exchange performance in a large-scale UF/DF vessel operated without active agitation during diafiltration.

3D-printed chromatography columns provide advantages over conventional packed bed resins, in particular, control over the multiscale structure that can be tailored to the needs of the separation. However, possible disparities between design and fabricated columns lessens this benefit. We apply 3D imaging to evaluate and compare printed structures to original CAD files to refine the design process, creating more effective separation capabilities for emergent biological products.

In vitro transcription enables large-scale mRNA production but generates difficult-to-remove double-stranded RNA (dsRNA) impurities. Existing chromatographic approaches for dsRNA removal are limited by scalability and manufacturing cost. We report a mixed-mode PrimaS monolith chromatography method that selectively clears dsRNA from 1–10 kb mRNA under RNA-compatible conditions, preserving integrity. Up to eightfold clearance of linear and structured dsRNA was achieved, supporting its potential as a platform purification solution.

This presentation introduces Hydrophobic Interaction Chromatography (HIC) as a powerful, aqueous alternative to traditional anion exchange and reverse-phase methods for ASO purification. We demonstrate how retaining the 5' hydrophobic blocking group enables a high dynamic binding capacity, allowing for an efficient bind-and-elute process. Our data highlights a systematic optimization of salt types and stepwise gradients to achieve >90% product yield with superior resolution of failure sequences and process-related impurities.

This talk shares an enhanced pre-use sanitization strategy for AVB Sepharose resin in adeno-associated virus (AAV) production. By evaluating various alcohol, acid, and NaOH combinations against environmental microbial contaminants and assessing the identified combination in AAV resin re-use studies, we offer a solution that enhances microbial control without compromising resin performance or product quality.

The presented dataset characterizes the impact of affinity chromatography process conditions on quality attributes of AAVs. We can thereby provide information to support a decision for or against direct loading after harvest versus prior concentration by TFF and add to the general understanding of AAVs as a product and the potential influence of proess conditions on yield losses, aggregation, and transduction efficiency.

We have developed a scalable, enzyme-free, closed workflow using fibrous chromatographic media before tangential flow filtration (TFF) to selectively remove extracellular chromatin (DNA/histones) from extracellular vesicle (EV) preparations. This strategy prevents proinflammatory responses while preserving EV yield and identity. Moreover, the use of fibrous chromatographic media improves purity, safety, and manufacturability, de-risking late-stage development and enabling GMP-compliant, large-scale EV production.

In biologics manufacturing, product-related impurities must be managed through a comprehensive strategy aligned with ICH Q6B guidelines. The emergence of new biologic modalities, such as bispecific antibodies, transpacific antibodies, and DVDs, has increased the complexity of impurity control due to the unique molecular structures and expanded impurity spectrum. This presentation focuses on low molecular weight species and discusses control strategies developed through downstream purification process optimization for those novel modalities.

We present the evaluation and optimization of MabSelect SuRe 70 for capture purification of a bispecific antibody in comparison with an alternative Protein A resin. MabSelect SuRe 70 demonstrated higher dynamic binding capacity, improved specific product recovery, and increased monomer purity. Further process improvements were achieved through optimization of elution conditions to enhance resolution and host cell protein (HCP) clearance.

This talk discusses purification process development strategies for a novel trispecific antibody (tsAb) intended for tumor therapy. It will highlight approaches to optimize downstream processing and support efficient, scalable manufacturing of complex antibody formats.

Hexameric IgGs are a class of complex biologics that offers unique pharmacokinetics and pharmacodynamic properties due to their high valency (up to 12 antigen binding sites), molecular weight (~900 kDa), or hydrodynamic radius. Despite their attractive profile for certain therapeutic indications, technical challenges in purification and analytics hinder their broader application. This talk aims to demystify some of the challenges by providing comprehensive molecule analysis/illustration and a couple of successful examples where key problems are identified and resolved. It would be of some guidance or introduction for those who are interested in making large and complex biologics for drug discovery and development.