Over the last few years we have continued development of the use of focused ion beams in biology for site-specific imaging of the interior of cellular and tissue specimens at spatial resolutions over an order of magnitude better than those currently achieved with optical microscopy. The principle of imaging is based on using a focused ion beam to create a cut at a designated site in the specimen, followed by viewing the newly generated surface with a scanning electron beam. Iteration of these two steps several times thus results in the generation of a series of surface maps of the specimen at regularly spaced intervals, which can be converted into a three-dimensional map of the specimen. We have extended the application of this method to a variety of eukaryotic cells and tissues to establish this as a powerful tool for cellular and sub-cellular imaging in 3D for biomedical and clinical applications. Highlights of progress over the last year include: (i) development of tools for correlative light and 3D electron microscopy, the first such demonstration showing that the same cells can be imaged first by fluorescence microscopy, and followed by imaging of the entire 3D volume by electron microscopy at 3D resolutions that are now better than 10 nm;(ii) development of """"""""atom probe tomography"""""""", a new technology for high resolution subcellular imaging and demonstrating its first application to biology by imaging mammalian cells at nanometer resolution and (iii) development of powerful tools for automation of data collection in focused ion beam microscopy in a collaboration with Fibics Inc. and its application to studying a variety of problems related to cell-cell communication on bacterial and viral infection.
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