Size Exclusion Chromatography Method Migration Between ACQUITY™ Premier and Arc™ Premier Systems

Significance of Topic

Size Exclusion Chromatography (SEC) is essential for the analysis of protein-based therapeutics, enabling detection and quantification of aggregation and fragmentation variants that impact drug efficacy and safety. Advances in column surface engineering have minimized undesired secondary interactions, improving accuracy and reproducibility in biopharmaceutical monitoring.

Objectives and Study Overview

This study aimed to demonstrate the successful migration of an SEC method for monoclonal antibodies from the ACQUITY Premier UPLC system to the Arc Premier UHPLC platform. The migration leveraged MaxPeak Premier Protein SEC columns with High Performance Surfaces (HPS) technology to maintain equivalent chromatographic performance across different particle sizes and system formats.

Methodology

An isocratic mobile phase of sterile filtered 1X PBS at pH 7.4 was employed at a defined flow rate and injection volume on a 1.7 µm ACQUITY Premier Protein SEC 250Å Column. Method parameters were scaled using the Waters Column Calculator (v2.0) to accommodate a 2.5 µm XBridge Premier Protein SEC 250Å Column on the Arc Premier system, ensuring consistent column loading and elution profiles.

Instrumentation

• ACQUITY Premier UPLC System
• Arc Premier UHPLC System
• ACQUITY Premier Protein SEC 250Å Column, 1.7 µm (4.6×300 mm)
• XBridge Premier Protein SEC 250Å Column, 2.5 µm (7.8×300 mm)
• Empower Chromatography Data System for data acquisition and custom field calculations
• Waters Column Calculator version 2.0 for method scaling

Main Results and Discussion

Analysis of Remicade and the biosimilar Renflexis on both systems yielded relative peak area differences of approximately 0.1% for aggregates and fragments. Resolution between monomer and low molecular weight species remained consistent, with slightly lower valley heights observed on the ACQUITY Premier due to smaller particle size. The minor resolution improvement did not affect overall purity assessments (Remicade: 99.4% vs 99.3%; Renflexis: 98.8% vs 98.7%). Additionally, throughput could be increased by ~20% on the 1.7 µm column by adjusting flow rate to match resolution on the 2.5 µm column.

Benefits and Practical Applications

• Reproducible separation of protein size variants with minimal secondary interactions
• Seamless method transfer between LC platforms supports pharmaceutical development, process monitoring, and QC release
• Flexible column formats and particle sizes accommodate diverse throughput and resolution requirements
• Simplified buffer conditions without high salt or organic modifiers enable biorelevant analysis

Future Trends and Opportunities

• Integration of rapid SEC workflows with automated DOE approaches for method optimization
• Expansion of HPS technology to additional column chemistries and particle sizes
• Coupling SEC platforms with orthogonal detectors (e.g., multi-angle light scattering) for comprehensive characterization
• Implementation of AI-driven data analysis to further enhance method transfer efficiency and predictive performance

Conclusion

This study confirms that SEC methods developed on one LC platform can be reliably migrated to another system while retaining equivalent performance. The combination of MaxPeak Premier HPS technology, column scaling tools, and optimized instrumentation ensures consistent, high-resolution analysis of monoclonal antibody size variants across diverse analytical environments.

References

1. Yang H, Koza SM, Yu YQ. Determination of Hydrodynamic Radius With MaxPeak Premier Protein SEC Columns. Waters Application Note, 720007625, 2022.
2. Kizekai L, Shiner SJ, Lauber MA. Waters ACQUITY and XBridge Premier Protein SEC 250Å Columns: A New Benchmark in Inert SEC Column Design. Waters Application Note, 720007493, 2022.