Abstract
We identify N-linked glycoproteins and deduce their spatial localization using spatially-resolved glycoproteomic guided by MALDI mass spectrometry N-glycan imaging on formalin-fixed paraffin embedded (FFPE) canine high-grade glioma tissues. The glycan HexNAc4-Hex5-NeuAc2 was found to be predominantly expressed in necrotic regions, whereas high mannose HexNAc2-Hex5 was predominantly found in benign regions. To determine the underlying sialo-glycoprotein, various regions in adjacent tissue sections were analysed. Haptoglobin is the protein associated with HexNAc4-Hex5-NeuAc2. Additionally, we identified several high-mannose (Hex2-HexNAc5) glycopeptides enriched in benign regions. This is the first report that directly links glycan imaging with intact glycopeptide identification. Over 400 N-glycosylated, O-GalNAcylated, O mannosylated, and S- and O- GlcNAcylated glycopeptides from over 30 proteins were identified. While N-glycosylation and O- mannosylation were similar between benign and tumor/necrotic sections, S- and O-GlcNAc glycopeptides were significantly decreased in tumor/necrotic sections, whereas sialylated O-GalNAc glycopeptides were significantly upregulated.
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