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Profiling and quantifying endogenous molecules in single cells using nano-DESI MS
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
2017 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 142, no 19, p. 3639-3647Article in journal (Refereed) Published
Abstract [en]

Molecular profiling of single cells has the potential to significantly advance our understanding of cell function and cellular processes of importance to health and disease. In particular, small molecules with rapid turn-over rates can reveal activated metabolic pathways resulting from an altered chemical environment or cellular events such as differentiation. Consequently, techniques for quantitative metabolite detection acquired in a higher throughput manner are needed to characterize the biological variability between seemingly homogenous cells. Here, we show that nanospray desorption electrospray ionization (nano-DESI) mass spectrometry ( MS) enables sensitive molecular profiling and quantification of endogenous species in single cells in a higher throughput manner. Specifically, we show a large number of detected amino acids and phospholipids, including plasmalogens, readily detected from single cheek cells. Further, by incorporating a phosphatidylcholine ( PC) internal standard into the nano-DESI solvent, we determined the total amount of PC in one cell to be 1.2 pmoles. Finally, we describe a higher throughput approach where molecules in single cells are automatically profiled. These developments in single cell analysis provide a basis for future studies to understand cellular processes related to drug effects, cell differentiation and altered chemical microenvironments.

Place, publisher, year, edition, pages
ROYAL SOC CHEMISTRY , 2017. Vol. 142, no 19, p. 3639-3647
National Category
Chemical Sciences
Identifiers
URN: urn:nbn:se:uu:diva-336430DOI: 10.1039/c7an00885fISI: 000411703800013PubMedID: 28835951OAI: oai:DiVA.org:uu-336430DiVA, id: diva2:1166526
Funder
Swedish Research Council, VR 621-2013-4231Swedish Foundation for Strategic Research , SSF ICA-6Available from: 2017-12-15 Created: 2017-12-15 Last updated: 2018-04-05Bibliographically approved
In thesis
1. Applications of nanospray desorption electrospray ionization mass spectrometry: In situ lipid and metabolite analysis from cells to tissue
Open this publication in new window or tab >>Applications of nanospray desorption electrospray ionization mass spectrometry: In situ lipid and metabolite analysis from cells to tissue
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Ambient mass spectrometry (MS) has proved to be an important addition to the bioanalytical toolbox. These methods perform analyte sampling and ionization under atmospheric pressure, and require very little sample preparation other than the sampling process in front of the machine. Nanospray desorption electrospray ionization (nano-DESI) is an ambient MS technique developed in 2010 that utilizes localized liquid extraction for surface sampling. The aim of this thesis was to explore the possibilities of this technique, and identify areas in which nano-DESI MS could further contribute to the community of MS-based surface analysis.

One such area was found to be mass spectrometry imaging (MSI) of small-molecule neurotransmitters. By the use of deuterated standards of acetylcholine, γ-aminobutyric acid and glutamate, the respective endogenous compounds were successfully imaged in coronal sections of rat brain. The use of internal standards was shown to be essential to compensatee for matrix effects in different regions of the brain. In a second imaging study, nano-DESI MSI was used to compare the chemical profiles of diabetic rat kidney tissue and control. Analysis was performed on kidney two weeks after diabetic onset, before any pathohistological changes relating to diabetic nephropathy can be seen in a microscope. In our study, it was shown that a large number of chemical species related to energy metabolism were detected with altered signal intensity in diabetic kidney tissue.

To push the limits of nano-DESI analysis, its use for single-cell analysis was evaluated. By placing buccal epithelial cells in contact with the nano-DESI probe, it was possible to identify 46 endogenous compounds and detect differences between cells from three human donors. In addition, it was shown that molecules from single cells on a surface could be detected by scanning the surface with the nano-DESI probe, which opens up for development of an automated analysis with higher throughput.

The last study in this thesis was concerned with method development rather than application, as it presented a setup for pneumatically assisted nano-DESI. Evaluation showed that the setup provided improved sensitivity in the analysis of small metabolites, and provided the possibility of using pure water as nano-DESI solvent.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. p. 78
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1660
Keywords
Mass spectrometry, mass spectrometry imaging (MSI), nanospray desorption electrospray ionization (nano-DESI), single-cell analysis, neurotransmitter imaging, diabetic nephropathy, pneumatic nebulization, lipidomics, metabolomics
National Category
Chemical Sciences
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-347674 (URN)978-91-513-0307-9 (ISBN)
Public defence
2018-05-25, A1:107a, BMC, Husargatan 3, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2018-05-03 Created: 2018-04-05 Last updated: 2018-10-08

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Bergman, Hilde-MarleneAndersson, Ingela

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