Advances in three-dimensional secondary ion mass spectrometry (SIMS) imaging have enabled visualizing the subcellular distributions of various lipid species within individual cells. However, the difficulty of locating organelles using SIMS limits efforts to study their lipid compositions. Here, the authors have assessed whether endoplasmic reticulum (ER)-Tracker Blue White DPX, which is a commercially available stain for visualizing the endoplasmic reticulum using fluorescence microscopy, produces distinctive ions that can be used to locate the endoplasmic reticulum using SIMS. Time-of-flight-SIMS tandem mass spectrometry (MS) imaging was used to identify positively and negatively charged ions produced by the ER-Tracker stain. Then, these ions were used to localize the stain and thus the endoplasmic reticulum, within individual human embryonic kidney cells that contained higher numbers of endoplasmic reticulum-plasma membrane junctions on their surfaces. By performing MS imaging of selected ions in parallel with the precursor ion (MS) imaging, the authors detected a chemical interference native to the cell at the same nominal mass as the pentafluorophenyl fragment from the ER-Tracker stain. Nonetheless, the fluorine secondary ions produced by the ER-Tracker stain provided a distinctive signal that enabled locating the endoplasmic reticulum using SIMS. This simple strategy for visualizing the endoplasmic reticulum in individual cells using SIMS could be combined with existing SIMS methodologies for imaging intracellular lipid distribution and to study the lipid composition within the endoplasmic reticulum.
Various lipid species and cholesterol form the selectively permeable membranes that surround the mammalian cell and compartmentalize its interior into organelles. Studies that employ cellular fractionation and biochemical analysis suggest that each organelle contains a distinct lipid composition, and this composition is related to organellar function. Advances in secondary ion mass spectrometry (SIMS) has enabled imaging the distributions of distinct lipid species on the surfaces of cells, and visualizing their intracellular distributions in three dimensions (3D) [1-3]. Organelle-specific labels that can be used to identify specific organelles with SIMS are needed in order to more accurately determine the lipid composition within individual organelles.
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