Focused issue on X-ray microscopy of biological materials.
The study of biological structures at different scales and resolution levels requires the use of imaging methods that are appropriate for each level of analysis. As recently reviewed in this journal (Volume 172, Issue 2 (2010)), the use of multiple, correlated methods is a powerful way to extend the analytical properties of each individual technique, normally resulting in more informative insights by overcoming the various limitations of the individual methods. In this context, there is an increasing demand for more fully quantitative approaches to image cells in a near to native state, a demand that is hardly satisfied by presently available methods. The emerging possibilities related to the use of X-rays to image intact, hydrated biological material thus present interesting new tools. The high penetrating power of X-rays, combined with recent advances in X-ray optics, have led to a number of very interesting results in three-dimensional analysis of biological samples. This focused issue aims to bring together a number of recent advances in the use of X-ray microscopy for structural analysis and chemical mapping of biological materials. This is a good moment to highlight work in this emerging field, as the use of soft X-rays and Fresnel zone plates now result in microscopes showing resolutions up to 20 nm. This special issue includes reports on the use of such instruments for three-dimensional imaging, using tomographic reconstructions of frozen cells in a fully hydrated state. Comparison of these tomographic reconstructions with data obtained by electron microscopy allows one to estimate both the possibilities and the limitations of x-ray based microscopy in the determination of cellular features. The possibility to detect different viral forms within eukaryotic cells also highlights the resolution level presently attainable using cryo X-ray tomography. The use of correlative approaches, combining, for example, the use of fluorescent probes for light microscopy with the X-ray microscopy analysis of frozen samples, has proven to be a powerful way to select the appropriate cells to image in further detail. As an example, light microscopy can be used to detect fluorophoretagged viral proteins within the mammalian cell environment, and X-ray microscopy can then be used to characterize virus-induced structures in the same cells. Another application of soft X-rays employs quantitative measurement of the absorption of X-rays, which is used to obtain, at the same time, cell morphology and a detailed estimation of the content of a cellular component (hemoglobin) in the host infected by a large protozoan cell at different states of its life cycle.