Improving N-glycan coverage using HPLC-MS with electrospray ionization at subambient pressure

Area of Science:

• Biochemistry
• Analytical Chemistry
• Biomarker Discovery

Background:

• Human serum glycan profiling using mass spectrometry (MS) shows promise for clinical biomarker discovery.
• Challenges in comprehensive profiling include low glycan ionization efficiency, large dynamic concentration ranges, and problematic fragmentation of large glycans.

Purpose of the Study:

• To investigate the utility of subambient pressure ionization with nanoelectrospray mass spectrometry (SPIN-MS) for expanding human serum glycome profiling.
• To compare SPIN-MS with conventional atmospheric pressure electrospray ionization mass spectrometry (ESI-MS) for enhanced glycan detection.

Main Methods:

• Liquid chromatographic separation on graphite columns was performed.
• Subambient pressure ionization with nanoelectrospray mass spectrometry (SPIN-MS) was utilized.
• Comparison was made with atmospheric pressure electrospray ionization mass spectrometry (ESI-MS) using a heated capillary inlet.

Main Results:

• SPIN-MS expanded the serum glycome profile compared to conventional ESI-MS.
• Higher charge states were observed for approximately half of the annotated glycans using SPIN-MS.
• 34 out of 130 detected glycans were exclusively identified using SPIN-MS, improving coverage of glycan families and larger glycans.

Conclusions:

• SPIN-MS significantly improves the coverage and depth of human serum glycan profiling.
• This enhanced profiling capability holds potential for advancing clinical biomarker discovery through more comprehensive glycomics analysis.