SUMMARY
Imaging filters developed over the last few years permit rapid elemental mapping by energy‐filtering transmission electron microscopy (EFTEM), with resolution and sensitivity limited primarily by the sample and by the TEM. We explore the attainable spatial resolution in the elemental maps theoretically and experimentally, and suggest optimized set‐up procedures for maximizing the resolution. The chromatic aberration of the objective lens of the microscope is shown to be a major limit. Its influence can be minimized by using small energy intervals and limited collection angles, but this is done at the cost of decreased collection efficiency. Resolution of better than 1 nm and sensitivities to less than a monolayer of elements with favourable edges are readily attainable in elemental maps obtained with acquisition times of 40 s total and less. Resolution better than 0·5 nm should be attainable with further optimization of the acquisition parameters.
SummaryWe illustrate the combined use of cryo-electron tomography and spectroscopic difference imaging in the study of subcellular structure and subcellular bodies in whole bacteria. We limited our goal and focus to bodies with a distinct elemental composition that was in a sufficiently high concentration to provide the necessary signal-to-noise level at the relatively large sample thicknesses of the intact cell. This combination proved very powerful, as demonstrated by the identification of a phosphorus-rich body in Caulobacter crescentus . We also confirmed the presence of a body rich in carbon, demonstrated that these two types of bodies are readily recognized and distinguished from each other, and provided, for the first time to our knowledge, structural information about them in their intact state. In addition, we also showed the presence of a similar type of phosphorus-rich body in Deinococcus grandis , a member of a completely unrelated bacteria genus. Cryo-electron microscopy and tomography allowed the study of the biogenesis and morphology of these bodies at resolutions better than 10 nm, whereas spectroscopic difference imaging provided a direct identification of their chemical composition.
Imaging filters produce energy-selected images in a few seconds, and chemical maps formed by processing of several images taken at different energy losses in typically less than one minute. On the other hand, imaging filters do not provide detailed spectra from each specimen point, and are vulnerable to artifacts due to variations in specimen thickness, and other effects influencing EELS background extrapolation and subtraction. These include diffraction contrast arising particularly in crystalline samples, edge overlap, and extended fine structures (EXELFS) in the pre-edge region caused by major edges at lower energies. We have therefore been exploring the practical usefulness of imaging filters on a range of specimens from materials science and biology. The results suggest that the imaging capability combined with full paralleldetection EELS performance delivers a very powerful experimental set-up.Figure 1 shows an energy-filtered bright field image of a steel sample containing about 1 % Cu, obtained at 120 keV with the Gatan Imaging Filter (GIF) attached to a Philips CM12ST microscope.
Abstract. 2014 An algorithm for automatic detection and identification of edges in an EELS spectrum is presented. It has the following features: 1) it compresses the dynamic range of EELS spectra and enhances the ionization edge signals via difference transforms, 2) it removes residual background, thereby isolating sharp features associated with the edge thresholds and noise, 3) it distinguishes true edge-threshold features from noise via statistical analysis. In addition to paving the way for rapid, automated EELS elemental analysis, the algorithm is capable of detecting edges which are easily overlooked by human analysts.
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