Removing Formaldehyde‐Induced Peptidyl Crosslinks Enables Mass Spectrometry Imaging of Peptide Hormone Distributions from Formalin‐Fixed Paraffin‐Embedded Tissues

Abstract

Linking molecular and chemical changes to human disease states depends on the availability of appropriate clinical samples, mostly preserved as formalin‐fixed paraffin‐embedded (FFPE) specimens stored in tissue banks. Mass spectrometry imaging (MSI) enables the visualization of the spatiotemporal distribution of molecules in biological samples. However, MSI is not effective for imaging FFPE tissues because of the chemical modifications of analytes, including complex crosslinking between nucleophilic moieties. Here we used an MS‐compatible inorganic nucleophile, hydroxylamine hydrochloride, to chemically reverse inter‐ and intra‐crosslinks from endogenous molecules. The analyte restoration appears specific for formaldehyde‐reactive amino acids. This approach enabled the MSI‐assisted localization of pancreatic peptides expressed in the alpha, beta, and gamma cells. Pancreatic islet‐like distributions of islet hormones were observed in human FFPE tissues preserved for more than five years, demonstrating that samples from biobanks can effectively be investigated with MSI.

Reproducible Lipid Alterations in Patient-Derived Breast Cancer Xenograft FFPE Tissue Identified with MALDI MSI for Pre-Clinical and Clinical Application

“…However, analysis of lipids has proven difficult in this type of tissue due to their depletion during initial processing with organic solvents, which is further compounded by the remaining lipids being implicated in cross-binding with other structures in the tissue [ 12 , 13 , 14 ]. For this reason, protocols that are able to remove, or reverse, the cross-links and liberate these trapped biomolecules are currently being developed [ 15 ]. Among these, it was recently shown that the inclusion of an antigen retrieval step in the MALDI-MSI sample preparation protocol increased the number of certain solvent-resistant lipids, particularly phosphatidylcholine (PC), sphingomyelin (SM), and phosphatidylserine (PS) species, that can be detected [ 16 ].…”