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. 2023 Oct 16;415(28):7011–7024. doi: 10.1007/s00216-023-04983-2

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — Combination of MALDI-MSI and MALDI-IHC on Human Breast Cancer Tissue. A An overview of the workflows performed. B Example H&E staining of two separate sections of the breast tissue; before workflow is started (left) and after workflows are completed (right). C Two sets of AmberGen MALDI-IHC images from slides processed by MALDI-MSI first (left tissue) and IMC first (right tissue). Markers visualized and their mass tags are labeled below images. MALDI-IHC step size was 20 µm. D Comparisons of two N-glycan ion images (1809.643 and 2174.763 m/z, 40 µm) from MALDI-MSI first and MALDI-IHC first along with (E) the whole spectra of N-glycans, (F) mean percent change when compared to MALDI-MSI first, and (G) the 10 most intense N-glycan peaks, ordered by MALDI-IHC first. H Images of collagen peptides with masses 843.394 m/z and 1034.572 m/z are compared for MALDI-MSI first and MALDI-IHC first. Step size of 40 µm was used. I The spectra of extracellular molecules are compared. J The percent difference in peak intensity displayed for a subset of selected peaks with respect to MALDI-MSI first. K Top 10 collagen peaks of highest intensity in MALDI-IHC first are compared