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Peer-reviewedCC BY-NC-ND 3.0Journal of visualized experiments : JoVE

Single-Cell Mass Spectrometry Imaging Using Desorption Electrospray Ionization Coupled To Orbitrap Mass Spectrometers

  • Nathan Colwell1,2
  • Dan Chen2
  • Deepti Bhusal1,2
  • Zongkai Peng2
  • Zhibo Yang1,2
  1. 1Department of Chemistry and Biochemistry, University of Oklahoma
  2. 2Department of Biochemistry and Physiology, University of Oklahoma Health Campus
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Abstract

Desorption electrospray ionization–mass spectrometry imaging (DESI–MSI) enables ambient, matrix-free molecular imaging with minimal sample preparation. This protocol describes a workflow for single-cell MSI of OVCAR-8 cells using DESI coupled to Orbitrap mass spectrometers. The system integrates a DESI sprayer, a modified mass spectrometer interface, a motorized XYZ stage, and an optical breadboard for high-spatial-resolution imaging. Cells are cultured on gridded glass coverslips, washed with ammonium formate solution to reduce salt interference, dried, and analyzed under ambient conditions. Solvent is delivered continuously using a nano-liquid chromatography system, and nebulizing nitrogen gas is regulated to maintain spray stability. The protocol also describes raster imaging, pixel control, and optical-to-mass spectrometry image registration for single-cell localization. High mass resolution and mass accuracy provided by Orbitrap detection enable differentiation of closely related molecular species and support single-cell metabolomic analysis. This protocol provides an adaptable strategy for implementing high-spatial-resolution single-cell MSI using multiple Orbitrap mass spectrometer platforms.

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Cite

BibTeXRIS
BibTeX
@article{colwell2026singlecell,
  title = {{Single-Cell Mass Spectrometry Imaging Using Desorption Electrospray Ionization Coupled To Orbitrap Mass Spectrometers}},
  author = {Nathan Colwell and Dan Chen and Deepti Bhusal and Zongkai Peng and Zhibo Yang},
  journal = {Journal of visualized experiments : JoVE},
  year = {2026},
  month = jun,
  doi = {10.3791/71691},
  url = {https://doi.org/10.3791/71691},
  note = {CC BY-NC-ND 3.0},
}
RIS
TY  - JOUR
TI  - Single-Cell Mass Spectrometry Imaging Using Desorption Electrospray Ionization Coupled To Orbitrap Mass Spectrometers
AU  - Nathan Colwell
AU  - Dan Chen
AU  - Deepti Bhusal
AU  - Zongkai Peng
AU  - Zhibo Yang
PY  - 2026
DA  - 2026/06/12
JO  - Journal of visualized experiments : JoVE
DO  - 10.3791/71691
UR  - https://doi.org/10.3791/71691
AB  - Desorption electrospray ionization–mass spectrometry imaging (DESI–MSI) enables ambient, matrix-free molecular imaging with minimal sample preparation. This protocol describes a workflow for single-cell MSI of OVCAR-8 cells using DESI coupled to Orbitrap mass spectrometers. The system integrates a DESI sprayer, a modified mass spectrometer interface, a motorized XYZ stage, and an optical breadboard for high-spatial-resolution imaging. Cells are cultured on gridded glass coverslips, washed with ammonium formate solution to reduce salt interference, dried, and analyzed under ambient conditions. Solvent is delivered continuously using a nano-liquid chromatography system, and nebulizing nitrogen gas is regulated to maintain spray stability. The protocol also describes raster imaging, pixel control, and optical-to-mass spectrometry image registration for single-cell localization. High mass resolution and mass accuracy provided by Orbitrap detection enable differentiation of closely related molecular species and support single-cell metabolomic analysis. This protocol provides an adaptable strategy for implementing high-spatial-resolution single-cell MSI using multiple Orbitrap mass spectrometer platforms.
SN  - 1940-087X
ER  -