A controlled raster of this highly focused primary beam across the sample surface can therefore, with proper standard materials as reference, provide quantitative images of chemical or isotope ratios with a lateral resolution permitting structures larger than a few hundred nanometers to be clearly resolved. The unique strength of the NanoSIMS ion microprobe is the ability to focus a relatively weak primary beam (order of pA either Cs + or O −) onto an extremely small spot, on the order of 100 nm, on the sample surface. In the multi-collector, the arrivals of up to seven specific isotopes are counted in parallel by ion detectors either electron multipliers or Faraday cups. The secondary ions are then transmitted through the magnetic sector mass spectrometer that resolves specific isotopes from isobaric interferences in the mass spectrum. The impacts of these primary ions create a collision cascade in the upper atomic layers of the sample, which leads to the ejection of secondary ions (either single atoms or small molecules) from the sample surface. Technically speaking, the NanoSIMS instrument is a double-focusing, magnetic sector, multi-collection ion microprobe operating in dynamic mode, i.e., with a continuous “primary” ion-beam bombarding a flat sample surface. The NanoSIMS ion microprobe instrument is an ultra-high-resolution isotope microscope that produces quantifiable maps of chemical and isotopic variations in a wide range of materials with a lateral resolution of around 100 nm. TeaserĬryoNanoSIMS: subcellular mapping of chemical and isotopic compositions of biological tissues in their most pristine post-mortem state. This opens new horizons in the study of fundamental processes at the tissue- and (sub)cellular level. With a cryo-workflow that includes vitrification by high pressure freezing, cryo-planing of the sample surface, and cryo-SEM imaging, the CryoNanoSIMS enables correlative ultrastructure and isotopic or elemental imaging of biological tissues in their most pristine post-mortem state. This capability is illustrated with nitrogen isotope as well as trace element mapping of freshwater hydrozoan Green Hydra tissue following uptake of 15N-enriched ammonium. Here, we report the development of a CryoNanoSIMS instrument that can perform isotope imaging of both positive and negative secondary ions from flat block-face surfaces of vitrified biological tissues with a mass- and image resolution comparable to that of a conventional NanoSIMS. To overcome these limitations an entirely cryogenic sample preparation and imaging workflow is required. However, the associated sample preparation methods all result in some degree of tissue morphology distortion and loss of soluble compounds. The development of nanoscale secondary ion mass spectrometry (NanoSIMS) has revolutionized the study of biological tissues by enabling, e.g., the visualization and quantification of metabolic processes at subcellular length scales.
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