Spatial distribution of isobaric lipids using high-resolution ion mobility with the DESI XS

Significance of the Topic

Desorption electrospray ionization imaging mass spectrometry (DESI-MS) enables direct chemical mapping of tissue surfaces under ambient conditions, offering valuable insights into spatial lipid distributions. However, overlapping m/z signals arising from diverse lipid structures limit specificity. High-resolution ion mobility, particularly cyclic multi-pass separation, addresses this challenge by discriminating ions based on structural and mobility differences, thereby improving specificity in lipid imaging.

Objectives and Study Overview

This study aimed to apply multi-pass high-resolution cyclic ion mobility separation within a DESI-MS workflow to enhance specificity in lipid imaging. The research focused on demonstrating the ability to resolve and spatially map isobaric phospholipids in tissue sections, comparing single-pass and multi-pass mobility separations.

Methods and Instrumentation

• Desorption Electrospray Ionization: DESI-XS source (Waters) using MeOH:H2O (98:2) with 0.1% formic acid at 2 µL/min; spray voltage 0.65 kV; nebulizing gas at 0.7 bar.
  
• Mass Spectrometer: Select Series Cyclic IMS quadrupole time-of-flight (ToF) with integrated cyclic ion mobility device (100 cm RF ion guide, >600 electrodes, multi-pass capability, up to 100 000 mass resolution).
  
• Sample Preparation: Fresh tissue sections thaw-mounted on glass slides and vacuum-dried; no additional sample prep.
  
• Data Acquisition: Survey imaging (m/z 50–1 200) using single and seven-pass cyclic IMS; drift time and m/z recorded.
  
• Data Processing: MassLynx V4.2, DriftScope V2.9, and HDI 1.6 for mobility analysis and image visualization.

Key Results and Discussion

• Single-pass IMS provided partial lipid separation but failed to resolve all isobaric species within complex phospholipid regions.
  
• Seven-pass cyclic IMS achieved enhanced resolution (~150), fully separating ions at m/z 810.6 into two drift time peaks (92.93 ms and 94.24 ms), corresponding to protonated PC(18:0_20:4) and sodiated PC(18:0_18:1) differing by 2.4 mDa.
  
• Tandem MS after mobility separation yielded distinct fragmentation patterns for each isobaric lipid, enabling confident structural assignments.
  
• Spatial mapping of separated species revealed unique localization patterns within mouse brain tissue, illustrating biological relevance of improved separation.

Benefits and Practical Applications

• Enhanced specificity in lipid imaging through discrimination of isobaric species.
  
• Direct tissue analysis without chromatography or extensive sample prep.
  
• Accurate molecular identification aided by post-mobility tandem MS.
  
• Potential applications in biomarker discovery, pathology, and metabolic studies.

Future Trends and Applications

• Integration of higher-pass IMS cycles and IMSⁿ workflows for deeper structural elucidation.
  
• Coupling mobility-enhanced DESI-MS with machine learning for automated tissue classification.
  
• Expansion to other molecular classes (metabolites, peptides) and clinical diagnostics.
  
• Development of compact, high-throughput instruments for routine spatial omics in research and QA/QC environments.

Conclusion

Multi-pass cyclic ion mobility separation significantly improves the specificity of DESI-MS lipid imaging by resolving isobaric species that single-pass methods cannot distinguish. The combination of high-resolution mobility separation and tandem MS provides robust molecular identification and spatial localization, advancing the capabilities of ambient tissue imaging.

References

Shrestha B., Towers M., Olivos H., Midey A., Claude E., Waters Corporation Poster, 2021.