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

• Published in: Biotechnology progress 2015-05, Vol.31 (3), p.840-848
• Main Authors: Kim, Kyoung-Jin, Kim, Yoon-Woo, Kim, Yun-Gon, Park, Hae-Min, Jin, Jang Mi, Hwan Kim, Young, Yang, Yung-Hun, Kyu Lee, Jun, Chung, Junho, Lee, Sun-Gu, Saghatelian, Alan
• Format: Article
• Language: English
• Subjects: Animals; Case-Control Studies; Cattle; comparative quantitation; Evaluation Studies as Topic; Fetuins - chemistry; Glycomics - methods; Glycosylation; Humans; Isotope Labeling; MALDI-MS; Male; N-Acetylneuraminic Acid; N-glycan; ortho-Aminobenzoates - chemistry; Polysaccharides - blood; Prostatic Neoplasms - blood; Reproducibility of Results; sialic acid neutralization; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; stable isotopic labeling; Up-Regulation
• ISSN: 8756-7938; 1520-6033
• DOI: 10.1002/btpr.2078

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 13C6/12C6‐2‐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 (R2 > 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. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:840–848, 2015