Abstract
We report the development of a 3D OrbiSIMS instrument for label-free biomedical imaging. It combines the high spatial resolution of secondary ion mass spectrometry (SIMS; under 200 nm for inorganic species and under 2 μm for biomolecules) with the high mass-resolving power of an Orbitrap (>240,000 at m/z 200). This allows exogenous and endogenous metabolites to be visualized in 3D with subcellular resolution. We imaged the distribution of neurotransmitters-gamma-aminobutyric acid, dopamine and serotonin-with high spectroscopic confidence in the mouse hippocampus. We also putatively annotated and mapped the subcellular localization of 29 sulfoglycosphingolipids and 45 glycerophospholipids, and we confirmed lipid identities with tandem mass spectrometry. We demonstrated single-cell metabolomic profiling using rat alveolar macrophage cells incubated with different concentrations of the drug amiodarone, and we observed that the upregulation of phospholipid species and cholesterol is correlated with the accumulation of amiodarone.
Highlights
As highlighted recently there are numerous analytical challenges associated with measuring the metabolome at the single-cell level.[2]
We report the development of a 3D OrbiSIMS instrument for label-free biomedical imaging
The 3D OrbiSIMS instrument We first describe the utility of the 3D OrbiSIMS from the perspective of a method for metabolic imaging and provide technical instrument details for interested readers
Summary
As highlighted recently there are numerous analytical challenges associated with measuring the metabolome at the single-cell level.[2]. Secondary ion mass spectrometry (SIMS) is an increasingly popular method in the life-sciences[11,12] since it provides high spatial resolution using a focused ion beam as the probe and image in 3D with ~5 nm depth resolution.[13,14] There have been significant developments to improve its application for biological studies, in particular to include MS/MS capability for identification Such developments include a quadrupole-orthogonal time-of-flight (ToF) spectrometer[15], the J105 (IONOPTIKA, UK) using a novel ToF-ToF design for continuous (non-pulsed) large cluster ion beams more suited to the detection of large biomolecules[12] and recently the addition of a linear ToF to a TRIFT (Physical Electronics, MN, U.S.A.) spectrometer allowing parallel MS/MS imaging.[16]. The signal-to-noise ratio, even for the abundant cholesterol molecule, was low despite the large pixel size and very long pixel acquisition times of 15 s
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