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Correlative organelle fluorescence microscopy and synchrotron X-ray chemical element imaging in single cells
- Roudeau, Stéphane, Carmona, Asuncion, Perrin, Laura, Ortega, Richard
- Analytical and bioanalytical chemistry 2014 v.406 no.27 pp. 6979-6991
- X-radiation, chemical speciation, fluorescence, fluorescence microscopy, fluorescent dyes, fluorescent proteins, image analysis, organelles, trace elements
- X-ray chemical element imaging has the potential to enable fundamental breakthroughs in the understanding of biological systems because chemical element interactions with organelles can be studied at the sub-cellular level. What is the distribution of trace metals in cells? Do some elements accumulate within sub-cellular organelles? What are the chemical species of the elements in these organelles? These are some of the fundamental questions that can be addressed by use of X-ray chemical element imaging with synchrotron radiation beams. For precise location of the distribution of the elements, identification of cellular organelles is required; this can be achieved, after appropriate labelling, by use of fluorescence microscopy. As will be discussed, this approach imposes some limitations on sample preparation. For example, standard immunolabelling procedures strongly modify the distribution of the elements in cells as a result of the chemical fixation and permeabilization steps. Organelle location can, however, be performed, by use of a variety of specific fluorescent dyes or fluorescent proteins, on living cells before cryogenic fixation, enabling preservation of element distribution. This article reviews the methods used for fluorescent organelle labelling and X-ray chemical element imaging and speciation of single cells. Selected cases from our work and from other research groups are presented to illustrate the potential of the combination of the two techniques.