SMASH Imaging: A Serial Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Strategy for High-Resolution...

Area of Science:

• Spatial biology
• Lipidomics
• Mass spectrometry imaging

Background:

• Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is crucial for spatial lipidomics, offering high sensitivity and resolution.
• Dual-polarity analysis and MS/MS data are essential for comprehensive lipid coverage and accurate annotation due to ionization preferences and limited structural information.
• Serial MALDI-MSI for multimodal data integration requires evaluation for feasibility at high spatial resolutions.

Purpose of the Study:

• To present SMASH imaging, a novel method for dual-polarity and MS2 spatial lipidomics within a single tissue section.
• To evaluate the feasibility of serial MALDI-MSI for high-resolution spatial lipidomics.
• To establish criteria for matrix compounds and assess the number of imaging layers for optimal data acquisition.

Main Methods:

• Developed SMASH imaging integrating dual-polarity and MS2 acquisition in one tissue section.
• Utilized specific matrix compounds (2,5-dihydroxyacetophenone and trans-2-[3-(4-tert-butylphenyl)-2-methyl-2-propenylidene]malononitrile) for dual-polarity analysis.
• Determined optimal SMASH imaging layers using coefficient of determination (R²) and evaluated performance at 30 μm and 5 μm resolutions.

Main Results:

• SMASH imaging visualized over 400 lipid species at 30 μm resolution with strict annotation criteria (S/N ≥ 50, m/z ≤ 10 mDa, CCS ≤ 20 Ų).
• Four layers of SMASH imaging enabled MS2-based characterization of 18 lipid species in mouse brain.
• Eight layers of SMASH imaging at 5 μm resolution annotated 25 lipid species, supported by spatial correlation metrics.

Conclusions:

• SMASH imaging provides a feasible approach for multimodal spatial lipidomics on a single tissue section.
• The method enhances lipidome coverage, annotation accuracy, and spatial resolution, enabling detailed lipid atlas creation.
• This technique facilitates comprehensive molecular characterization and spatial mapping of lipids in biological tissues.