MALDI mass spectrometry imaging on formalin-fixed paraffin-embedded tissue: A spoonful of sugar of the <em>N-</em>glycome  — ASN Events
  1. Schedule
  2. Symposium Five: Glycoproteomics and Glycomics
  3. MALDI mass spectrometry imaging on formalin-fixed paraffin-embedded tissue: A spoonful of sugar of the N-glycome 

MALDI mass spectrometry imaging on formalin-fixed paraffin-embedded tissue: A spoonful of sugar of the N-glycome  (#021)

Matthew Briggs 1 2 , Arun Dass 3 , Julia Kuliwaba 4 5 , Dzenita Muratovic 4 5 , David Findlay 4 , Martin Oehler 6 , Nicki Packer 3 , Peter Hoffmann 1 2
  1. University of Adelaide, Adelaide, S.A., Australia
  2. Adelaide Proteomics Centre, School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA
  3. Faculty of Science, Biomolecular Frontiers Research Centre, Macquarie University, Sydney, NSW
  4. Discipline of Orthopaedics and Trauma, The University of Adelaide, Adelaide, SA, Australia
  5. Bone and Joint Research Laboratory, SA Pathology, Adelaide, SA, Australia
  6. Department of Gynaecological Oncology, Royal Adelaide Hospital, Adelaide, SA, Australia

In humans, it is estimated that 50-60% of proteins are glycosylated. N-linked glycans are of particular interest in diseases such as ovarian cancer and osteoarthritis because structural alterations have been observed. Further investigation of these alterations may unearth novel biomarkers for early stage diagnosis. A novel method for investigating tissue-specific N-linked glycans was recently developed by our group on formalin-fixed paraffin-embedded (FFPE) murine kidney. Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) spatially profiles N-glycans in tissue-specific regions, while through liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) the corresponding glycol compositions are structurally characterized.

Regions of interest such as tumour, stroma, adipose and necrotic tissue were isolated from FFPE serous ovarian cancer tissue sections while cartilage and bone marrow tissue were isolated from FFPE osteoarthritic bone tissue sections. N- and O-linked glycans were structurally characterized through enzymatic peptide-N-glycosidase F (PNGase F) release of N-glycans, followed by β-elimination of O-glycans. The released glycans were analyzed through porous graphitized carbon liquid chromatography (PGC-LC) and collision induced negative mode fragmentation analysis. Following structural characterization, high resolution MALDI-IMS revealed the spatial distribution of these identified N-glycans and their tissue-specificity on consecutive FFPE tissue sections. For example, tumour and non-tumour tissue regions established clear demarcation based on their N-glycan distribution.   

MALDI-IMS and LC-ESI-MS/MS were used as complementary techniques to generate high resolution images and structural information of tissue-specific N-glycans. Furthermore, O-glycoforms were characterized for the first time in FFPE ovarian tumour tissue sections. Application of this method enabled the first steps in the glycomic quest for the Holy Grail: an early stage diagnosis biomarker.