Multiplexed imaging mass spectrometry of the extracellular matrix using serial enzyme digests from formalin-fixed paraffin-embedded tissue sections
- 18 November 2020
- journal article
- research article
- Published by Springer Science and Business Media LLC in Analytical and Bioanalytical Chemistry
- Vol. 413 (10), 2709-2719
- https://doi.org/10.1007/s00216-020-03047-z
Abstract
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. Graphical AbstractKeywords
Funding Information
- National Center for Advancing Translational Sciences (UL1 TR000445)
- Center for Strategic Scientific Initiatives, National Cancer Institute (R21CA240148)
- National Heart, Lung, and Blood Institute (HL007260)
- American Heart Association (16GRNT31380005)
- South Carolina Center of Economic Excellence
- National Institute of General Medical Sciences (P20 GM103542)
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