Refined on-tissue chemical derivatization expands MALDI-MSI capabilities for spatial metabolomics of mammalian organs and metastatic tumors using EDC and 4-APEBA
4 affiliations
- 1Leibniz-Institut für Analytische Wissenschaften—ISAS—e.V, Otto-Hahn-Str. 6b, 44227 Dortmund, Germany
- 2Department for Dermatology, Venerology and Allergology, Research Center One Health, University Hospital Essen, German Cancer Consortium, Partner Site Essen & University Alliance Ruhr, NCT West, Campus Essen, 45147 Essen, Germany
- 3Lipidomics, Faculty of Chemistry, University of Duisburg-Essen, 45141 Essen, Germany
- 4Medizinische Fakultät, Medizinische Proteom-Center (MPC), Ruhr-Universität Bochum, 44801 Bochum, Germany
Abstract
Background: Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is widely used in metabolomics because it enables high-resolution spatial characterization of molecular distributions directly from tissue. Nevertheless, comprehensive analysis of small, polar metabolites remains difficult due to limited ionization efficiencies and diverse chemical structures. On-tissue chemical derivatization (OTCD) has emerged as an effective strategy to improve ionization efficiency, sensitivity, and selectivity for these analytes.
Methods: In this study, we established and systematically optimized an OTCD approach for spatial analysis of carbonyl- and carboxyl-containing metabolites in murine tissues using 4-APEBA in combination with carbodiimide crosslinker EDC. Reagent concentrations, solvent composition, and spray parameters were optimized to achieve reproducible derivatization across multiple tissue types.
Results: The optimized protocol enabled reproducible and confident detection of carbonyl- and carboxyl-containing metabolites in positive ion mode through the characteristic bromine isotopic signature of 4-APEBA. Application to metastatic liver and lung tissue demonstrated the method’s performance in biologically complex environments. Pronounced metabolic heterogeneity correlated with histopathological features in tumor-rich liver sections, and clear tumor-specific localizations were obtained in lung tissue with only small metastatic foci. Metabolic differences between tumor and adjacent healthy regions were consistent with established cancer metabolism pathways, including glycolytic reprogramming.
Conclusion: This study is the first to optimize and investigate sample preparation conditions for mouse tissue application of a reliable OTCD workflow using EDC and 4-APEBA for MALDI-MSI-based spatial metabolomics, with localized OTCD-MSI of melanoma metastases. By utilizing this combination of reagents, carbonyl and carboxyl moiety containing molecules are confidently annotated based on isotopic signatures in positive mode ionization. The application is capable of interrogating metabolic heterogeneity in clinically relevant tumor models and demonstrates its value for future investigations of tumor metabolism.
Graphical Abstract:
Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1186/s12935-026-04494-x.
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BibTeX
@article{fecke2026refined,
title = {{Refined on-tissue chemical derivatization expands MALDI-MSI capabilities for spatial metabolomics of mammalian organs and metastatic tumors using EDC and 4-APEBA}},
author = {Antonia Fecke and Luiza Martins Nascentes Melo and Ann-Cathrin Groba and Gabriele Allies and Feyza Cansiz and Siva Swapna Kasarla and Sven Heiles and Albert Sickmann and Alpaslan Tasdogan and Karl W. Smith},
journal = {Cancer Cell International},
year = {2026},
month = nov,
doi = {10.1186/s12935-026-04494-x},
url = {https://doi.org/10.1186/s12935-026-04494-x},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Refined on-tissue chemical derivatization expands MALDI-MSI capabilities for spatial metabolomics of mammalian organs and metastatic tumors using EDC and 4-APEBA
AU - Antonia Fecke
AU - Luiza Martins Nascentes Melo
AU - Ann-Cathrin Groba
AU - Gabriele Allies
AU - Feyza Cansiz
AU - Siva Swapna Kasarla
AU - Sven Heiles
AU - Albert Sickmann
AU - Alpaslan Tasdogan
AU - Karl W. Smith
PY - 2026
DA - 2026/11/04
JO - Cancer Cell International
DO - 10.1186/s12935-026-04494-x
UR - https://doi.org/10.1186/s12935-026-04494-x
AB - Background: Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is widely used in metabolomics because it enables high-resolution spatial characterization of molecular distributions directly from tissue. Nevertheless, comprehensive analysis of small, polar metabolites remains difficult due to limited ionization efficiencies and diverse chemical structures. On-tissue chemical derivatization (OTCD) has emerged as an effective strategy to improve ionization efficiency, sensitivity, and selectivity for these analytes. Methods: In this study, we established and systematically optimized an OTCD approach for spatial analysis of carbonyl- and carboxyl-containing metabolites in murine tissues using 4-APEBA in combination with carbodiimide crosslinker EDC. Reagent concentrations, solvent composition, and spray parameters were optimized to achieve reproducible derivatization across multiple tissue types. Results: The optimized protocol enabled reproducible and confident detection of carbonyl- and carboxyl-containing metabolites in positive ion mode through the characteristic bromine isotopic signature of 4-APEBA. Application to metastatic liver and lung tissue demonstrated the method’s performance in biologically complex environments. Pronounced metabolic heterogeneity correlated with histopathological features in tumor-rich liver sections, and clear tumor-specific localizations were obtained in lung tissue with only small metastatic foci. Metabolic differences between tumor and adjacent healthy regions were consistent with established cancer metabolism pathways, including glycolytic reprogramming. Conclusion: This study is the first to optimize and investigate sample preparation conditions for mouse tissue application of a reliable OTCD workflow using EDC and 4-APEBA for MALDI-MSI-based spatial metabolomics, with localized OTCD-MSI of melanoma metastases. By utilizing this combination of reagents, carbonyl and carboxyl moiety containing molecules are confidently annotated based on isotopic signatures in positive mode ionization. The application is capable of interrogating metabolic heterogeneity in clinically relevant tumor models and demonstrates its value for future investigations of tumor metabolism. Graphical Abstract: Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1186/s12935-026-04494-x.
SN - 1475-2867
ER -