Multimodal Chemical Analysis of the Brain by High Mass Resolution Mass Spectrometry and Infrared Spectroscopic Imaging

Abstract

The brain functions through chemical interactions between many different cell types, including neurons and glia. Acquiring comprehensive information on complex, heterogeneous systems requires multiple analytical tools, each of which have unique chemical specificity and spatial resolution. Multimodal imaging generates complementary chemical information via spatially localized molecular maps, ideally from the same sample, but requires method enhancements that span from data acquisition to interpretation. We devised a protocol for performing matrix-assisted laser desorption/ionization (MALDI)-Fourier transform ion cyclotron resonance-mass spectrometry imaging (MSI), followed by infrared (IR) spectroscopic imaging on the same specimen. Multimodal measurements from the same tissue provide precise spatial alignment between modalities, enabling more advanced image processing such as image fusion and sharpening. Performing MSI first produces higher quality data from each technique compared to performing IR imaging before MSI. The difference is likely due to fixing the tissue section during MALDI matrix removal, thereby preventing analyte degradation occurring during IR imaging from an unfixed specimen. Leveraging the unique capabilities of each modality, we utilized pan sharpening of MS (mass spectrometry) ion images with selected bands from IR spectroscopy and midlevel data fusion. In comparison to sharpening with histological images, pan sharpening can employ a plethora of IR bands, producing sharpened MS images while retaining the fidelity of the initial ion images. Using Laplacian pyramid sharpening, we determine the localization of several lipids present within the hippocampus with high mass accuracy at 5 μm pixel widths. Further, through midlevel data fusion of the imaging data sets combined with k-means clustering, the combined data set discriminates between additional anatomical structures unrecognized by the individual imaging approaches. Significant differences between molecular ion abundances are detected between relevant structures within the hippocampus, such as the CA1 and CA3 regions. Our methodology provides high quality multiplex and multimodal chemical imaging of the same tissue sample, enabling more advanced data processing and analysis routines.

Lipid Heterogeneity between Astrocytes and Neurons Revealed by Single‐Cell MALDI‐MS Combined with Immunocytochemical Classification

“…90 %l ower) as well ap olymer contaminant (Supporting Information, Figure S1). Thus,w ed ecided to perform ICC after MALDI-MS ( Figure 1a-c) in amanner similar to that described in our recent paper in which the coupling of vibrational spectroscopy and MALDI-MS imaging of tissue slices was reported; [15] using MALDI-MS first, we observed high-quality mass spectra of molecules,i ncluding small metabolites and lipids. Fort his procedure to succeed, the MALDI matrix (e.g., 2,5dihydroxybenzoic acid), which had been thoroughly cocrystalized with the cells,h ad to be removed while ensuring that the cells remained anchored to the same location on the slide.Finally,the antibody must still recognize its epitope.W e found that paraformaldehyde was effective in both fixing the cells and removing the matrix ( Figure S2 g).…”

Exploring the Fundamental Structures of Life: Non‐Targeted, Chemical Analysis of Single Cells and Subcellular Structures

“…[125] We expect multimodal measurements to remain ar apidly developing approach for increasing chemical coverage. [126] By coupling single-cell MS techniques with complementary targeted and non-targeted techniques,s uch as single-cell transcriptomics,s pectroscopy,a nd staining,e ven more information can be garnered.…”

High Spatial Resolution Ambient Ionization Mass Spectrometry Imaging Using Microscopy Image Fusion Determines Tumor Margins

“…Mass spectrometry imaging (MSI) provides spatially resolved chemical information of the surface of biological samples in a on tissue sections 37,38,39 . Meanwhile, these MSI have to be operated under high vacuum conditions and may limit their compatibility with conventional imaging methods for pathological examinations.…”