title: Comparative glycomic profiling of isotopically permethylated N-glycans by liquid chromatography/electrospray ionization mass spectrometry.
abstract: BACKGROUND
  Mass spectrometry based comparative glycomics is essential for disease biomarker discovery. However, developing a reliable quantification method is still a challenging task.
  
  
  METHODS
  We here report an isotopic labeling strategy employing stable isotopic iodomethane for comparative glycomic profiling by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS). N-Glycans released from model glycoproteins and blood serum samples were permethylated with iodomethane ('light') and iodomethane-d1 or -d3 ('heavy') reagents. Permethylated samples were then mixed at equal volumes prior to LC/ESI-MS analysis.
  
  
  RESULTS
  Peak intensity ratios of N-glycans isotopically permethylated (Heavy/Light, H/L) were almost equal to the theoretical values. Observed differences were mainly related to the purity of 'heavy' iodomethane reagents (iodomethane-d1 or -d3). The data suggested the efficacy of this strategy to simultaneously quantify N-glycans derived from biological samples representing different cohorts. Accordingly, this strategy is effective in comparing multiple samples in a single LC/ESI-MS analysis. The potential of this strategy for defining glycomic differences in blood serum samples representing different esophageal diseases was explored.
  
  
  CONCLUSIONS
  LC/ESI-MS comparative glycomic profiling of isotopically permethylated N-glycans derived from biological samples and glycoproteins reliably defined glycan changes associated with biological conditions or glycoproteins expression. As a biological application, this strategy permitted the reliable quantification of glycomic changes associated with different esophageal diseases, including high grade dysplasia, Barrett's disease, and esophageal adenocarcinoma.
authors.affiliation: Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409, USA.
authors.forename: Yunli
authors.initials: Y
authors.lastname: Hu
authors.forename: Janie L
authors.initials: JL
authors.lastname: Desantos-Garcia
authors.forename: Yehia
authors.initials: Y
authors.lastname: Mechref
mesh_terms: D001471:Barrett Esophagus / Q000097:blood; D002236:Carbohydrate Conformation; D016022:Case-Control Studies; D002853:Chromatography, Liquid / Q000379:methods*; D004938:Esophageal Neoplasms / Q000097:blood; D054794:Glycomics / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis / Q000097:blood / Q000737:chemistry; D006801:Humans; D008745:Methylation; D008956:Models, Chemical; D011134:Polysaccharides / Q000032:analysis* / Q000097:blood / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D006023:Glycoproteins; D011134:Polysaccharides