What can PREMIER do for your glycan analysis?

Importance of the Topic

Glycan profiling is essential in biopharmaceutical development and quality control to ensure proper glycosylation patterns that affect efficacy and safety.

Study Aims and Overview

This study evaluates the performance of ACQUITY PREMIER Glycan BEH Amide Columns featuring MaxPeak high performance surfaces and fetuin passivation, comparing them to conventional BEH Amide columns for glycan separation reliability and recovery.

Methodology and Instrumentation

• Columns: ACQUITY PREMIER Glycan BEH Amide, 1.7 µm, 130 Å, with MaxPeak surfaces; conventional BEH Amide 1.7 µm for comparison
• Passivation: Fetuin coating applied to minimize analyte–surface interactions
• Analytes: RapiFluor-MS Glycan Performance Test Standard, including highly sialylated N-glycans and a phosphorylated glycan (M7-1P)
• Separation: HILIC mode on UPLC with fluorescence detection
• Measurements: Retention time repeatability across injections, peak area reproducibility, recovery assessment

Key Results and Discussion

• Retention Time Precision: PREMIER columns achieved 0.1–0.2% RSD versus 1.1–2.0% for conventional columns across six major sialylated glycan peaks
• Reproducibility: Overlay chromatograms demonstrated consistent peak shapes and intensities over four injections and after passivation
• Recovery: PREMIER columns exhibited improved recovery of the phosphorylated glycan M7-1P, with stable peak areas across injections
• Surface Performance: Fetuin passivation on PREMIER surfaces effectively reduced analyte losses compared to unpassivated conventional columns

Benefits and Practical Applications

• Enhanced sensitivity and reproducibility for complex glycan profiles
• Reduced sample loss securing higher data confidence in QC and research environments
• Robust performance for high-throughput glycomics workflows in pharmaceutical and academic laboratories

Future Trends and Opportunities

• Development of advanced surface chemistries for further analyte protection
• Integration with mass spectrometry for comprehensive glycoform analysis
• Expansion to glycopeptide and intact glycoprotein separations
• Automation and high-throughput platforms enabling large-scale glycan screening

Conclusion

The ACQUITY PREMIER Glycan BEH Amide Column with MaxPeak surfaces and fetuin passivation delivers superior precision, reproducibility, and analyte recovery compared to conventional columns, making it a reliable choice for advanced glycan analysis.

Reference

Waters Corporation Application Note 720007234EN April 2021