Measurement of Absolute Concentration at the Subcellular Scale

Abstract

The concentration of a pharmaceutical drug or bioactive metabolite within the target organelle influences the effects elicited by the drug or metabolite. Although the relative concentrations of many compounds of interest within subcellular compartments have been measured, measurements of absolute concentrations in the organelle remain elusive. In this Perspective, we discuss a significant advance in using nano secondary ion mass spectrometry (nanoSIMS) to measure the absolute concentration of a 13C-labeled metabolite within secretory vesicles, as reported by Thomen et al. in the April issue of ACS Nano.

Chemical Analysis of Single Cells and Organelles

“…In addition, the optimization of NanoSIMS parameters, such as the secondary ion emission steady state (Figure B), was discussed. This study was also thoroughly explored in a perspective by Kraft and co-workers …”

“…This study was also thoroughly explored in a perspective by Kraft and co-workers. 147 Together with high sensitivity and mass resolution, Nano-SIMS currently features a spatial resolution down to approximately 50 nm (lateral) and 10 nm (depth), and it is therefore on the way to becoming one of the most useful single-cell analysis methods in MSI. It also finds many applications in the biogeochemistry and marine environment fields, and it is used to study how single-cell and oligocellular organisms such as phytoplankton, algae, and bacteria metabolize and respond to environmental changes.…”

Depth Correction of 3D NanoSIMS Images Shows Intracellular Lipid and Cholesterol Distributions while Capturing the Effects of Differential Sputter Rate

“…To permit detecting a molecule of interest that lacks a distinctive elemental signature with NanoSIMS, that molecule may be labeled with a nonperturbing rare stable isotope that does not alter its native intracellular distribution or metabolism. − For example, the subcellular distributions of rare isotope-labeled cholesterol and sphingolipids have been revealed by NanoSIMS imaging. ,,− Also, drugs labeled with rare isotopes have been localized at their sites of action . Small (∼100 nm) intracellular vesicles containing rare isotope-labeled dopamine have been imaged with NanoSIMS, and a strategy was developed for quantifying the dopamine concentration within individual vesicles, providing insight into neurotransmitter release. ,− …”

Nanoprojectile Secondary Ion Mass Spectrometry for Analysis of Extracellular Vesicles

“…Secondary ion mass spectrometry (SIMS) is ideally suited for sensitive analysis of nanometer-sized objects or ultrathin layers. − Typically, nanoscale analysis with SIMS requires the use of a focused ion beam (a few 10 nm) comprised low energy (keV) atomic or polyatomic ions. − Unfortunately, impacts of atomic or polyatomic ions result in extensive fragmentation of analyte molecules, and nanoscale analysis is limited to elemental analysis, thus the investigation of biological materials requires the use of a rare isotope or elemental tag . Recently, Nolan and colleagues reported a nano-SIMS methodology where a focused ion beam probe was used to analyze a tissue section labeled with isotopically tagged Abs. , This strategy showed considerable promise but, with a spatial resolution of ∼100 nm, it may not be suitable for the analysis of single EVs.…”