ac500298a_si_001.pdf (118.41 kB)
Dual Modifications Strategy to Quantify Neutral and Sialylated N‑Glycans Simultaneously by MALDI-MS
journal contribution
posted on 2015-12-17, 02:57 authored by Hui Zhou, Peter G. Warren, John W. Froehlich, Richard S. LeeDifferences
in ionization efficiency among neutral and sialylated
glycans prevent direct quantitative comparison by their respective
mass spectrometric signals. To overcome this challenge, we developed
an integrated chemical strategy, Dual Reactions for Analytical Glycomics
(DRAG), to quantitatively compare neutral and sialylated glycans simultaneously
by MALDI-MS. Initially, two glycan samples to be compared undergo
reductive amination with 2-aminobenzoic acid and 2-13[C6]-aminobenzoic acid, respectively. The different isotope-incorporated
glycans are then combined and subjected to the methylamidation of
the sialic acid residues in one mixture, homogenizing the ionization
responses for all neutral and sialylated glycans. By this approach,
the expression change of relevant glycans between two samples is proportional
to the ratios of doublet signals with a static 6 Da mass difference
in MALDI-MS and the change in relative abundance of any glycan within
samples can also be determined. The strategy was chemically validated
using well-characterized N-glycans from bovine fetuin and IgG from
human serum. By comparing the N-glycomes from a first morning (AM)
versus an afternoon (PM) urine sample obtained from a single donor,
we further demonstrated the ability of DRAG strategy to measure subtle
quantitative differences in numerous urinary N-glycans.