Gel-assisted mass spectrometry imaging enables sub-micrometer spatial lipidomics

Chan, Yat Ho; Pathmasiri, Koralege C.; Pierre-Jacques, Dominick; Hibbard, Maddison C.; Tao, Nannan; Fischer, Joshua L.; Yang, Ethan; Cologna, Stephanie M.; Gao, Ruixuan

doi:10.1038/s41467-024-49384-w

Cited by 3 publications

References 41 publications

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Spatial proteomics of single cells and organelles on tissue slides using filter-aided expansion proteomics

Dong,

Jiang,

et al. 2024

Nat Commun

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Hydrogel-based tissue expansion combined with mass spectrometry (MS) offers an emerging spatial proteomics approach. Here, we present a filter-aided expansion proteomics (FAXP) strategy for spatial proteomics analysis of archived formalin-fixed paraffin-embedded (FFPE) specimens. Compared to our previous ProteomEx method, FAXP employed a customized tip device to enhance both the stability and throughput of sample preparation, thus guaranteeing the reproducibility and robustness of the workflow. FAXP achieved a 14.5-fold increase in volumetric resolution. It generated over 8 times higher peptide yield and a 255% rise in protein identifications while reducing sample preparation time by 50%. We also demonstrated the applicability of FAXP using human colorectal FFPE tissue samples. Furthermore, for the first time, we achieved bona fide single-subcellular proteomics under image guidance by integrating FAXP with laser capture microdissection.

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Spatial proteomics of single cells and organelles on tissue slides using filter-aided expansion proteomics

Dong,

Jiang,

et al. 2024

Nat Commun

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Examination of Lipid Distributions in Hydrogel-Expanded Mouse Brain Tissue Using Imaging Mass Spectrometry

Samuel,

Yan,

Liang

et al. 2024

Preprint

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Imaging is an essential tool in biological research, and imaging mass spectrometry uniquely provides a label-free approach with high molecular specificity. However, imaging mass spectrometry is limited in spatial resolution, which in turn limits the biological structures and processes that can be studied at small dimensions. Custom lens setups and altered optical paths have been used to shrink the diameter of the incident laser beam probe in matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry to achieve high spatial resolutions (< 5 μm). However, these research-grade instruments are complex and expensive, making high spatial resolution imaging experiments unrealistic for the broader community. An alternative method for improving spatial resolution is through physical magnification of the substrate, which has been well established in the subfield of expansion microscopy (ExM). ExM leverages superabsorbent hydrogels for isotropic expansion of tissues and retention of analytes containing fluorescent tags. While typical ExM involves covalently anchoring the analyte of interest to the hydrogel network, lipid retention without anchoring has been recently demonstrated for imaging mass spectrometry. Herein, we demonstrate expansion imaging mass spectrometry (ExIMS) of expanded, whole brain tissue and examine lipid distributions in both positive and negative ion mode across multiple brain structures. A linear expansion factor of 4.5-fold is achieved and used to obtain high spatial resolution images of mouse brain cerebellum. Approximately 95% of lipids in both positive and negative ion mode are retained in expanded tissue compared to unexpanded tissue. Additionally, the majority of lipid distributions across the brain are maintained post-expansion. Alterations to the hydrogel formulation (e.g., crosslinker density) can significantly affect the ability of ExIMS to maintain accurate lipid distributions in expanded tissue.

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Ten-Fold Expansion MALDI Mass Spectrometry Imaging of Tissues and Cells at 500 nm Resolution

Xie,

Wang,

Diao

et al. 2024

Preprint

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Achieving high spatial resolution in matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is crucial for detailed molecular mapping in biological tissues and cells. However, conventional MALDI-MSI platforms are typically limited to spatial resolutions of 5-20 μm, restricting their ability to visualize fine subcellular structures. Here, we present a novel ten-fold expansion MALDI-MSI (10X ExMSI) method that attains 500 nm spatial resolution without the need for specialized optics or customized instrumentation. By physically expanding biological samples using a swellable hydrogel matrix, our method maintains high retention and broadens the detection range of lipids, ensuring comprehensive lipid profiling. We demonstrated the effectiveness of 10X ExMSI by visualizing intricate subcellular structures within mouse brain tissues, including individual nuclei, astrocytes, Purkinje cells, and, notably, dendritic arborizations-features previously unresolvable using mass spectrometry imaging. Applying 10X ExMSI to single-cell analysis of cultured cells revealed detailed spatial distributions of lipids at the subcellular to organelle level. Notably, the method is fully compatible with standard commercial MALDI-MSI platforms, enabling widespread adoption without significant additional investment. The 10X ExMSI offers a transformative tool for high-resolution molecular imaging, opening new avenues for molecular analysis in diverse biological and biomedical fields, including neuroscience, pathology, and cellular biology.

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Gel-assisted mass spectrometry imaging enables sub-micrometer spatial lipidomics

Cited by 3 publications

References 41 publications

Spatial proteomics of single cells and organelles on tissue slides using filter-aided expansion proteomics

Spatial proteomics of single cells and organelles on tissue slides using filter-aided expansion proteomics

Examination of Lipid Distributions in Hydrogel-Expanded Mouse Brain Tissue Using Imaging Mass Spectrometry

Ten-Fold Expansion MALDI Mass Spectrometry Imaging of Tissues and Cells at 500 nm Resolution

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