Imaging mass spectrometry (IMS) is becoming an essential technique in lipidomics. Still, many questions remain open, precluding it from achieving its full potential. Among them, identification of species directly from the tissue is of paramount importance. However, it is not an easy task, due to the abundance and variety of lipid species, their numerous fragmentation pathways, and the formation of a significant number of adducts, both with the matrix and with the cations present in the tissue. Here, we explore the fragmentation pathways of 17 lipid classes, demonstrating that in-source fragmentation hampers identification of some lipid species. Then, we analyze what type of adducts each class is more prone to form. Finally, we use that information together with data from on-tissue MS/MS and MS 3 to refine the peak assignment in a real experiment over sections of human nevi, to demonstrate that statistical analysis of the data is significantly more robust if unwanted peaks due to fragmentation, matrix, and other species that only introduce noise in the analysis are excluded.

For many years, traditional histology has been the gold standard for the diagnosis of many diseases. However, alternative and powerful techniques have appeared in recent years that complement the information extracted from a tissue section. One of the most promising techniques is imaging mass spectrometry applied to lipidomics. Here, we demonstrate the capabilities of this technique to highlight the architectural features of the human kidney at a spatial resolution of 10 μm. Our data demonstrate that up to seven different segments of the nephron and the interstitial tissue can be readily identified in the sections according to their characteristic lipid fingerprints and that such fingerprints are maintained among different individuals ( n = 32). These results set the foundation for further studies on the metabolic bases of the diseases affecting the human kidney.

Sublimation is a widely used method for matrix deposition in imaging mass spectrometry experiments. Still, most of the time, standard glass sublimators are used for this purpose, which do not enable optimal matrix deposition reproducibility, compromising inter-experiment comparison of the results. Here, we present an in-house designed stainless steel sublimator in which the parameters that have the strongest influence over matrix deposition reproducibility can be easily monitored. Using sections of human colon biopsies, we demonstrate the capabilities of this new prototype.

Here, we present a simple and cost-effective procedure to improve the spatial resolution of the commercial MALDI source of a LTQ Orbitrap. Based in spatial filtering techniques, we demonstrate that, with minimal modifications of the original setup, the system resolution can be pushed forward to <10 μm. The improved system performance is demonstrated by means of MALDI imaging of human colon biopsies.

Ferroptosis, a form of regulated necrosis characterized by peroxidation of lipids such as arachidonic acid‐containing phosphatidylethanolamine (PE), contributes to the pathogenesis of acute kidney injury (AKI). We have characterized the kidney lipidome in an experimental nephrotoxic AKI induced in mice using folic acid and assessed the impact of the ferroptosis inhibitor Ferrostatin‐1. Matrix‐assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) was used to assess kidney lipidomics and it discriminated between glomeruli, medulla, and cortex in control kidneys, AKI kidneys, and AKI + Ferrostatin‐1 kidneys. Out of 139 lipid species from 16 classes identified, 29 (20.5%) showed significant differences between control and AKI at 48 h. Total PE and lyso‐sulfatide species decreased, while phosphatidylinositol (PI) species increased in AKI. Dysregulated mRNA levels for Pemt, Pgs1, Cdipt, and Tamm41, relevant to lipid metabolism, were in line with the lipid changes observed. Ferrostatin‐1 prevented AKI and some AKI‐associated changes in lipid levels, such as the decrease in PE and lyso‐sulfatide species, without changing the gene expression of lipid metabolism enzymes. In conclusion, changes in the kidney lipid composition during nephrotoxic AKI are associated with differential gene expression of lipid metabolism enzymes and are partially prevented by Ferrostatin‐1. © 2022 The Pathological Society of Great Britain and Ireland.