An ideal method for the analysis of N-glycans would both identify the isomeric structure and deliver a true picture of the relative, if not absolute, amounts of the various structures in one sample. Porous graphitic carbon chromatography coupled with electrospray ionization mass spectrometry (ESI-MS) detection has emerged as a method with a particularly high potential of resolving isomeric oligosaccharides, but little attention has so far been paid to quantitation of the results obtained. In this work, we isolated a range of structures from Man5 to complex type N-glycans with zero to four sialic acids and blended them into an equimolar “glyco tune mix”. When subjected to liquid chromatography–ESI-MS in positive and negative modes, the glyco tune mix clearly demonstrated the futility of quantitation of N-glycans of different overall composition, different number of sialic acids, and strongly differing size without compensation for their very different molar responses. Relative quantitation of human plasma N-glycans was performed with correction factors deduced from this external glyco tune mix. Addition of just one isotope-coded internal standard with enzymatically added 13C-galactose led to absolute quantification in the same experiment. [Figure not available: see fulltext.]
CITATION STYLE
Grünwald-Gruber, C., Thader, A., Maresch, D., Dalik, T., & Altmann, F. (2017). Determination of true ratios of different N-glycan structures in electrospray ionization mass spectrometry. Analytical and Bioanalytical Chemistry, 409(10), 2519–2530. https://doi.org/10.1007/s00216-017-0235-8
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