Enhanced Recovery of Released Glycans Using QuanRecovery Sample Vials with MaxPeak High Performance Surfaces (HPS) Technology

Importance of the Topic

Glycan analysis by hydrophilic interaction chromatography with fluorescence and mass spectrometric detection (HILIC-FLR/MS) is essential for characterizing critical quality attributes of biotherapeutics. However, low and variable recoveries of released glycans caused by nonspecific adsorption to sample vial surfaces can compromise data accuracy and reproducibility. Addressing this surface-induced sample loss at the vial level complements advances in LC fluidics and column technologies to ensure more robust glycan profiling workflows.

Study Goals and Overview

This application note investigates the impact of QuanRecovery Sample Vials featuring MaxPeak High Performance Surfaces (HPS) on the recovery and in-solution stability of challenging glycans. Using both sialylated and phosphorylated glycan standards, the study compares FLR and MS responses over multiple injections from standard polypropylene vials versus HPS-coated vials. The objective is to demonstrate improved assay accuracy and consistency when integrating QuanRecovery vials into existing HILIC-FLR/MS methods.

Methodology and Instrumentation

The study employed a BioAccord LC-MS System coupled with an ACQUITY Premier BEH Amide Glycan Column for HILIC separations and fluorescence detection. Glycan samples, including a Rapifluor-MS Glycan Performance Test Standard and mannose-6-phosphate (M6P)-bearing glycans from Cathepsin D, were reconstituted and split between standard polypropylene autosampler vials and QuanRecovery vials. Fifteen consecutive injections were performed for each vial type, monitoring peak areas by FLR for sialylated species and MS for phosphorylated species to assess recovery and signal consistency.

Main Results and Discussion

• Sialylated glycans (FA2BG2S2) exhibited nearly two-fold higher and significantly more consistent FLR signals when stored and injected from QuanRecovery vials compared with polypropylene vials.
• Low-abundance M6P glycans showed stable MS responses over 15 injections only when stored in QuanRecovery vials, indicating reduced analyte-surface interactions.
• The MaxPeak HPS coating effectively minimizes nonspecific adsorption of both charge-bearing and hydrophobic glycan species, enhancing in-solution stability during autosampler storage.

Benefits and Practical Applications

• Enhanced sample recovery leads to improved assay accuracy and reproducibility in glycan profiling workflows.
• Compatibility with existing HILIC-FLR/MS methods allows straightforward implementation without altering established protocols.
• Better analytical consistency supports critical quality attribute monitoring in biopharmaceutical development and manufacturing.

Future Trends and Opportunities

• Expanding HPS-coated vial technologies to other challenging biomolecules such as monoclonal antibodies and peptides to further reduce sample loss.
• Integrating surface-optimized consumables with automated sample preparation platforms for high-throughput glycomics.
• Developing standardized guidelines for assessing vial surface performance across diverse analytical modalities.

Conclusion

QuanRecovery Sample Vials with MaxPeak HPS Technology offer a simple and effective solution to mitigate nonspecific adsorption in HILIC-FLR/MS glycan analysis. By enhancing recovery and signal consistency for both sialylated and phosphorylated glycans, these vials complement advanced LC systems and columns, enabling more accurate, reproducible, and robust glycoprofiling methods critical for biotherapeutic quality control.

References

1. Liu X. and Lauber M. MaxPeak High Performance Surfaces Technology Improves HILIC Profiling of Released N-Glycans. Waters Application Note, 720007263EN, 2021.