The combination of stable isotope probing (SIP) and nanoSIMS has increasingly been used in microbiology field over the past 15 years. The SIP-nanoSIMS approach is highly sensitive and allows tracing and quantifying metabolic rates at the level of single-cells, based on the assimilation of isotopically-labelled substrates into the cell biomass. It allows us to explore biological interactions between uncultured microorganisms, especially in the context of spatial and trophic relationships in biological systems where exchanges of nutrients can be extremely rapid, e.g., symbiotic or syntrophic interactions. Generally, SIP-nanoSIMS can answer questions that are out of reach for conventional approaches, ranging from microbe-host and microbe-microbe interactions (including single-cell ecophysiology) to cell-cell nutrient or metabolite exchanges, and interactions between cells and their organic/inorganic matrix. However, the wider application of this approach in microbiology field is currently hampered by the necessity of custom made, sample dependent preparation work-flows for correct identification and quantification of cellular processes and interactions within a spatial context. During this presentation, Niculina Musat (Helmholtz Centre for Environmental Research, Germany) discussed a few case studies on the application of SIP-nanoSIMS to trace microbial processes and interactions, including:

1. tracing the flow of carbon in the rhizosphere
2. studying the capability of microbial colonizers of plastics to degrade antibiotics
3. tracing the nutrient and water flow between fungi and microbial cells

Together, these case studies illustrate the necessity of thoughtful consideration and design of suitable sample preparation as well as the necessity of using correlative analyses to add a new level of resolution to the microbial identity-function conundrum.

Cable bacteria are filamentous bacteria that evolved an ingenious division of labour in which redox transformations in distant cells are coupled via long-distance electron transport. Cells in deeper sediment layers oxidize hydrogen sulfide, the electrons generated are then transported via “wires” along the longitudinal axis of the filament towards cells residing in the oxic zone, where they are used to reduce oxygen. To investigate the cellular physiology and the interactions among cells within a filament, the metabolic activity of single cells was tracked by nanoSIMS after probing with different stable isotopes (¹³C, ¹⁵N and ¹⁸O). Nicole Geerlings (Utrecht University) showed how these SIP-nanoSIMS analyses provided important insights into the division of labour, growth, cell cycle, and polyphosphate dynamics in multicellular cable bacteria.

NanoSIMS is one of the few techniques suitable for the detection and quantification of volatiles and trace elements in natural and experimental samples at the micron scale. Laurent Remusat (National Museum of Natural History, France) presented two case to show the advances that nanoSIMS can bring to the understanding of crucial geological processes: element partitioning during planetary differentiation and crustal melting.

By Matthew Steinhauser (University of Pittsburg). The aim of this presentation is to present our experience coupling stable isotope tracer methodologies with NanoSIMS quantitative imaging in applications to biology and biomedical research. We have applied NanoSIMS imaging to study metabolic processes including glucose, amino acid, lipid, and nucleic acid metabolism and cell turnover in model organisms and humans at subcellular resolution. Key vignettes involving application to adipose tissue biology, cardiovascular biology, and cancer will demonstrate the breath of possibilities with this technology and underscore the power of obtaining quantitative functional measurements at the single and subcellular level.

By Mary Kraft (University of Illinois Urbana-Champaign). Strategies for imaging the distributions of metabolically labeled cholesterol and sphingolipids on and within mammalian cells with a NanoSIMS instrument will be presented. By combining metabolic label incorporation and NanoSIMS imaging, the distributions cholesterol and sphingolipids in the plasma membrane and within subcellular structures within mammalian cells have been visualized. A new computational tool for converting NanoSIMS depth profiling data into accurate three-dimensional images of intracellular component distribution without requiring cell sectioning or correlated sample analysis with complementary techniques will also be presented. NanoSIMS imaging of the distributions of membrane components on and within intact cells may enable a better understanding of the roles of membrane lipids and cholesterol in cellular function.

By Silvio Rizzoli (Georg-August University, Goettingen). Brain function is intimately bound to the function of the synapses, small contact sites between neurons that enable the transfer of information. Synapse dysfunction is at the root of most brain diseases, and therefore a holistic understanding of synapse function has been the “holy grail” of neuroscience for decades. Our laboratory analyzes synapses by combining advanced imaging techniques in the optical domain with conventional mass spectrometry, with electron microscopy and with nanoscale secondary ion mass spectrometry (nanoSIMS). Over the last decade, we could derive a thorough understanding of the dynamics of synapses over long time periods, in a fashion that could not be even approached without nanoSIMS imaging, and we also approached the SIMS-based analysis of small brain-regulating drugs. We conclude that nanoSIMS is a promising tool in the biomedical domain, which has been exploited far too little.