Imaging and 3D Elemental Characterization of Intact Bacterial Spores by High-Resolution Secondary Ion Mass Spectrometry

Abstract

We present a quantitative, imaging technique based on nanometer-scale secondary ion mass spectrometry for mapping the 3D elemental distribution present in an individual micrometer-sized Bacillus spore. We use depth profile analysis to access the 3D compositional information of an intact spore without the additional sample preparation steps (fixation, embedding, and sectioning) typically used to access substructural information in biological samples. The method is designed to ensure sample integrity for forensic characterization of Bacillus spores. The minimal sample preparation/alteration required in this methodology helps to preserve sample integrity. Furthermore, the technique affords elemental distribution information at the individual spore level with nanometer-scale spatial resolution and high (microg/g) analytical sensitivity. We use the technique to map the 3D elemental distribution present within Bacillus thuringiensis israelensis spores.

Direct chemical evidence for sphingolipid domains in the plasma membranes of fibroblasts

“…After calculating the moving average over a 3 × 3-pixel window, the lateral resolution of the enrichment image was 87 nm, compared with the 70-nm diameter of the analysis beam. Based on our operating conditions and the sputtering rate determined for biological samples (26), the sputtering depth (<2 nm) was much less than the plasma membrane's thickness (7.5 nm) (27), ensuring that a minimum number of secondary ions were collected from the underlying cytoplasm.…”

AFM and NanoSIMS analyses of Vaccinia virions

“…For this work, up to five species were collected simultaneously. Because the lateral resolution of the primary ion beam was at the scale of the virions, we used an imaging depth profile approach, which has the potential to provide higher spatial resolution in depth than could be achieved laterally [2]. This approach also avoided the need for sectioning.…”

Data Processing for 3D Mass Spectrometry Imaging

“…This information serves as a powerful tool for finding disease biomarkers as well as for understanding and developing drug delivery systems [25,26]. While 2D-MS imaging has been widely applied to the analysis of thin tissue sections, it has also been recognized that it is highly valuable to acquire 3D spatial distributions of the chemicals in a tissue volume or in an entire organ [27][28][29][30][31][32][33][34]. The two basic approaches used for 3D-MS imaging are, first, depth profiling using an ionization source that ablates tissue and second, recording a sequence of 2D images from serial sections taken from a tissue volume and then combining this information.…”

“…The two basic approaches used for 3D-MS imaging are, first, depth profiling using an ionization source that ablates tissue and second, recording a sequence of 2D images from serial sections taken from a tissue volume and then combining this information. In the depth profiling experiments, ablation of the tissue material is used to expose lower layers of tissue for analysis; this has been achieved with high energy ion beams in secondary ion mass spectrometry (SIMS) imaging [29,33,35] or with lasers in methods that include matrix assisted laser desorption (MALDI) [5,36,37], laser ablation electrospray ionization (LAESI) [28,38], and laser ablation followed by atmospher-ic-pressure afterglow (LA-FAPA) [39]. In the alternative serial-sectioning approach, a volume of tissue is sliced into thin sections and each of the sections is imaged using standard 2D-MS imaging.…”