METABOLOMIC/LIPIDOMIC DESI IMAGING OF DIFFERENT CELL CULTURES

Significance of the Topic

Lipidomics provides essential insights into the structural and functional roles of lipids in biological systems. DESI mass spectrometry imaging enables direct, label-free spatial mapping of lipid distribution in cell cultures without extensive preparation.

Objectives and Study Overview

This study investigates lipid distribution and localization in three cell lines: two gut epithelial models (Caco2, HT29-MTX) and a basophil leukemia line (RBL), using DESI-MSI in positive and negative ion modes to maximize lipid coverage.

Methodology and Sample Preparation

• Cell lines Caco2 and HT29-MTX cultured for 21 days; RBL cultured for 2–3 days on glass coverslips.
• Media replaced every 48 hours; live/dead staining performed to assess cell viability prior to imaging.
• Samples washed with 150 mM ammonium acetate, dried in a desiccator for 15 minutes, then mounted directly for DESI analysis.
• DESI parameters: 98:2 methanol:water spray at 1.5 µl/min, capillary voltage 4.5 kV, pixel sizes 20 and 50 µm at 80 and 100 µm/sec stage speeds.

Instrumentation Used

• Waters Xevo G2-XS Q-ToF mass spectrometer
• Desorption electrospray ionization (DESI) source
• MassLynx and High Definition Imaging (HDI) 1.5 software for data acquisition and visualization
• MetaboAnalyst online platform for multivariate statistical analysis

Main Results and Discussion

• Positive mode imaging revealed strong signals in the m/z 700–1,000 range, with m/z 798.54 (putative PC(34:1)K+) as the most abundant peak.
• Multivariate analysis (PCA, heatmaps) distinguished cell lines by lipid profiles: Caco2 and RBL exhibited broader lipid diversity, whereas HT29-MTX formed a tighter cluster.
• Triglycerides were detected only in Caco2 and HT29-MTX cultures, not in RBL.
• Negative mode highlighted oleic acid (m/z 281.25) as the dominant signal across all samples; PE species prevailed in Caco2 and RBL, while PI(36:1)H– was prominent in HT29-MTX.
• High-resolution imaging at 20 µm pixel size improved visualization of cell agglomerates and lipid co-localization.

Benefits and Practical Applications

• Direct, label-free lipid imaging with minimal sample preparation.
• Rapid data acquisition suitable for high-throughput analyses.
• High spatial resolution enables sub-cellular localization studies.
• Potential for cell line phenotyping and drug response assessment based on lipid signatures.

Future Trends and Applications

• Integration of DESI-MSI with tandem MS for definitive lipid identification.
• Advancements in spatial resolution approaching sub-cellular scales.
• Application of machine learning for automated lipidome classification.
• Extension to three-dimensional cell cultures and organotypic models.

Conclusion

In summary, DESI-MSI on the Xevo G2-XS platform offers a powerful, direct approach to map lipid distributions in cell cultures, enabling detailed phenotypic characterization and advancing analytical capabilities in lipidomics.

Reference

• MetaboAnalyst: https://www.metaboanalyst.ca/MetaboAnalyst/faces/home.xhtml