LC-MS Analytics in Bioprocessing: Automation-Driven Analysis of Product Quality Attributes and Nutrient Monitoring

Significance of the Topic

Early and accurate monitoring of product quality attributes (PQAs) and nutrient levels is vital in biopharmaceutical manufacturing to ensure safety, efficacy, and consistency of therapeutic proteins. Automation of LC-MS workflows accelerates sample preparation, reduces human error, and delivers critical data at earlier stages of process development, enabling faster decision-making and reducing time to market.

Objectives and Study Overview

This application note presents recent enhancements to the Waters Bioprocess Walk-Up Solutions with Automation Portal, aimed at empowering bioprocess teams with expanded, faster LC and LC-MS analytics. Key goals include:

• Streamlining sample handling and direct injection to LC or LC-MS.
• Implementing automated Protein A cleanup and vacuum filtration protocols.
• Updating software for comprehensive reporting of low/high molecular weight species and charge variants.
• Introducing a rapid nine-minute cell culture media method.

Methodology and Instrumentation

Automated workflows integrate the Andrew+ Pipetting Robot with the BioAccord LC-MS system via the ACQUITY Automation Portal. Protocols include magnetic bead–based Protein A purification, vacuum filtration using AcroPrep plates, plate sealing, and direct plate insertion.

Used Instrumentation

• Andrew+ Pipetting Robot with BenchHub, ARS, Extraction+ module, Plate Sealer+.
• BioAccord LC-MS System coupled to ACQUITY Premier BSM with Automation Portal.
• Intact Mass 1.8 and UNIFI 1.9.4 for MS data processing.
• Waters_connect 3.3 for chromatography reporting and OneLab 1.20.3 for protocol orchestration.

Main Results and Discussion

Automated Protein A and dilution workflows delivered reliable glycoform profiles and light-chain measurements. Filtration studies showed optimal debris removal using vacuum filtration at stepped pressures with predilution in PBS, preserving cell integrity. SEC-MS methods enabled clear separation and quantification of low/high molecular weight species with automated LMWS/HMWS reporting. Data export tools and an Excel visualization macro facilitated rapid comparison of bioreactor conditions and time courses across multiple metabolites and glycoforms.

Benefits and Practical Applications

• High throughput: seamless sample prep and direct LC-MS injection.
• Consistent results: reduced variability through automated sealing, filtration, and handling.
• Comprehensive reporting: glycoforms, aggregation metrics, and charge variants.
• Connected workflows: support for Ambr bioreactor integration and external data packages.

Future Trends and Opportunities

Advances in AI-driven data analysis, expanded metabolite and glycan libraries, and tighter integration with real-time process control systems will further accelerate bioprocess optimization. Emerging cloud-based informatics platforms and digital-twin modeling promise enhanced predictive capabilities and seamless cross-site standardization.

Conclusion

The updated Bioprocess Walk-Up Solutions simplify and accelerate LC-MS–based PQA monitoring, delivering faster, more reliable data with minimal manual intervention. These automation-driven workflows support critical cell line selection, process development, and quality control decisions, ultimately reducing development timelines and improving product consistency.

Reference

1. Alelyunas YA et al. Simplifying Bioreactor In-Process Monitoring with Waters Bioprocess Walk-Up Solutions. Application Note 720008062, 2023.
2. Alelyunas YA et al. Monitoring Intact Glycoprofiles and Spent Media Metabolites in Samples from Sartorius Ambr 250 High Throughput Bioreactor System. Application Note 720008042, 2023.
3. Alelyunas YA, Gray J, Wrona MD, Boyce P. Introducing a Rapid Throughput LC-MS Method for Cell Culture Media Nutrient and Metabolite Analysis. Application Note 720008170, 2024.
4. Shion H, Boyce P, Berger SJ, Yu YQ. INTACT Mass – Rapid Mass Confirmation and Purity Assessment of Biotherapeutics. Application Note 720007547, 2022.
5. Koza SM, Jiang AHW, Yu YQ. Rapid SEC-UV Analysis of Monoclonal Antibodies Using Ammonium Acetate Mobile Phases. Application Note 720007852, 2022.