Journal
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
Volume 413, Issue 10, Pages 2709-2719Publisher
SPRINGER HEIDELBERG
DOI: 10.1007/s00216-020-03047-z
Keywords
Multiplexed; Imaging mass spectrometry; Chondroitin sulfate; N-Glycans; Elastin; Collagen
Funding
- American Heart Association [16GRNT31380005]
- NIH/NIGMS [P20 GM103542]
- National Center for Advancing Translational Sciences [UL1 TR000445]
- NIH/NHLBI [HL007260]
- NCI/IMAT [1R21CA207779]
- South Carolina Centers of Economic Excellence SmartState program
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This study presents a multiplexed imaging mass spectrometry method to spatially localize and selectively access the extracellular matrix on formalin-fixed paraffin-embedded tissue sections. A novel serial enzyme strategy has been developed to define the extracellular matrix, including PTMs, from a single tissue section for MALDI-IMS applications to overcome the challenge of capturing complex ECM peptides.
We report a multiplexed imaging mass spectrometry method which spatially localizes and selectively accesses the extracellular matrix on formalin-fixed paraffin-embedded tissue sections. The extracellular matrix (ECM) consists of (1) fibrous proteins, post-translationally modified (PTM) via N- and O-linked glycosylation, as well as hydroxylation on prolines and lysines, and (2) glycosaminoglycan-decorated proteoglycans. Accessing all these components poses a unique analytical challenge. Conventional peptide analysis via trypsin inefficiently captures ECM peptides due to their low abundance, intra- and intermolecular cross-linking, and PTMs. In previous studies, we have developed matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI-IMS) techniques to capture collagen peptides via collagenase type III digestion, both alone and after N-glycan removal via PNGaseF digest. However, in fibrotic tissues, the buildup of ECM components other than collagen-type proteins, including elastin and glycosaminoglycans, limits efficacy of any single enzyme to access the complex ECM. Here, we have developed a novel serial enzyme strategy to define the extracellular matrix, including PTMs, from a single tissue section for MALDI-IMS applications.
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