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Biotechnol Prog. 2015 May-Jun;31(3):840-8. doi: 10.1002/btpr.2078. Epub 2015 Apr 15.

Stable isotopic labeling-based quantitative targeted glycomics (i-QTaG).

Author information

1
Dept. of Chemical Engineering, Soongsil University, Seoul, 156-743, Republic of Korea.
2
School of Chemical and Biological Engineering, Seoul National University, Seoul, 151-742, Republic of Korea.
3
Div. of Mass Spectrometry Research, Korea Basic Science Institute, Ochang, 363-883, Republic of Korea.
4
Dept. of Bio-Analytical Science, University of Science and Technology, Daejeon, 305-764, Republic of Korea.
5
Dept. of Microbial Engineering, College of Engineering, Konkuk University, Seoul, 143-701, Republic of Korea.
6
Dept. of Internal Medicine, Dongguk University Ilsan Hospital, College of Medicine, Dongguk University, Goyang, 401-773, Si Republic of Korea.
7
Dept. of Biochemistry and Molecular Biology and Cancer Research Institute, Seoul National University College of Medicine, Seoul, 110-799, Republic of Korea.
8
School of Chemical and Biomolecular Engineering, Pusan National University, Pusan, 609-735, Republic of Korea.
9
Clayton Foundations Laboratories for Peptide Biology, Salk Institute, La Jolla, CA, 92037.

Abstract

Mass spectrometry (MS) analysis combined with stable isotopic labeling is a promising method for the relative quantification of aberrant glycosylation in diseases and disorders. We developed a stable isotopic labeling-based quantitative targeted glycomics (i-QTaG) technique for the comparative and quantitative analysis of total N-glycans using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). We established the analytical procedure with the chemical derivatizations (i.e., sialic acid neutralization and stable isotopic labeling) of N-glycans using a model glycoprotein (bovine fetuin). Moreover, the i-QTaG using MALDI-TOF MS was evaluated with various molar ratios (1:1, 1:2, 1:5) of (13) C6 /(12) C6 -2-aminobenzoic acid-labeled glycans from normal human serum. Finally, this method was applied to direct comparison of the total N-glycan profiles between normal human sera (n = 8) and prostate cancer patient sera (n = 17). The intensities of the N-glycan peaks from i-QTaG method showed a good linearity (R(2) > 0.99) with the amount of the bovine fetuin glycoproteins. The ratios of relative intensity between the isotopically 2-AA labeled N-glycans were close to the theoretical molar ratios (1:1, 1:2, 1:5). We also demonstrated that the up-regulation of the Lewis antigen (~82%) in sera from prostate cancer patients. In this proof-of-concept study, we demonstrated that the i-QTaG method, which enables to achieve a reliable comparative quantitation of total N-glycans via MALDI-TOF MS analysis, has the potential to diagnose and monitor alterations in glycosylation associated with disease states or biotherapeutics.

KEYWORDS:

MALDI-MS; N-glycan; comparative quantitation; sialic acid neutralization; stable isotopic labeling

PMID:
25832445
DOI:
10.1002/btpr.2078
[Indexed for MEDLINE]

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