title: Imaging of N-linked glycans from formalin-fixed paraffin-embedded tissue sections using MALDI mass spectrometry.
abstract: Aberrant glycosylation is associated with most of the diseases. Direct imaging and profiling of N-glycans on tissue sections can reveal tissue-specific and/or disease-associated N-glycans, which not only could serve as molecular signatures for diagnosis but also shed light on the functional roles of these biomolecules. Mass spectrometry imaging (MSI) is a powerful tool that has been used to correlate peptides, proteins, lipids, and metabolites with their underlying histopathology in tissue sections. Here, we report an MSI technique for direct analysis of N-glycans from formalin-fixed paraffin-embedded (FFPE) tissues. This technique consists of sectioning FFPE tissues, deparaffinization, and rehydration of the sections, denaturing tissue proteins, releasing N-linked glycans from proteins by printing peptide-N-glycosidase F over the sections, spray-coating the tissue with matrix, and analyzing N-glycans by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Brain sections from a C57BL/6 mouse were imaged using this technique at a resolution of 100 μm. Forty-two N-glycans were analyzed from the mouse brain section. The mass spectrometry images were used to study the relative abundance of oligomannose, nonfucosylated, and fucosylated complex N-glycans in different brain areas including isocortex, hippocampal formation, and brainstem and specific glycans associated with different areas of the brain were identified. Furthermore, glioblastoma tumor xenografts in a NOD/SCID mouse were imaged. Several glycans with differential expression in tumor versus normal brain tissues were identified. The MSI technique allows for imaging of N-glycans directly from FFPE sections. This method can potentially identify tissue-specific and/or disease-associated glycans coexpressed with other molecular signatures or within certain histological structures.
authors.affiliation: Department of Biomedical Engineering, ‡Department of Pathology, School of Medicine, Johns Hopkins University , Baltimore, Maryland 21231, United States.
authors.forename: Shadi
authors.initials: S
authors.lastname: Toghi Eshghi
authors.forename: Shuang
authors.initials: S
authors.lastname: Yang
authors.forename: Xiangchun
authors.initials: X
authors.lastname: Wang
authors.forename: Punit
authors.initials: P
authors.lastname: Shah
authors.forename: Xingde
authors.initials: X
authors.lastname: Li
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism; D001932:Brain Neoplasms / Q000378:metabolism; D002236:Carbohydrate Conformation; D005557:Formaldehyde; D005909:Glioblastoma / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D008297:Male; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D057054:Molecular Imaging / Q000379:methods*; D016612:Paraffin Embedding; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000737:chemistry; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D016707:Tissue Fixation / Q000379:methods*; D023041:Xenograft Model Antitumor Assays
chemical_list: D011134:Polysaccharides; D005557:Formaldehyde; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase