Abstract
The need to determine the precise subcellular distribution of specific proteins and macromolecular complexes in cells and tissues has been the major driving force behind the development of new molecular-genetic and chemical-labeling approaches applicable to high-resolution, correlated, multidimensional microscopy. This short review is intended to provide an overview of recently developed and widely used electron microscopy (EM)-compatible probes, including tetracysteine tags, mini singlet oxygen generator (MiniSOG), time-specific tag for the age measurement of proteins (TimeSTAMP) with MiniSOG, and enhanced ascorbate peroxidase (APEX). We describe how these highly specific and genetically introduced EM probes are now used, in conjunction with lower resolution light microscopic methods, to obtain wide field or dynamic records of live preparation or of large maps in 3D using recently developed laboratory-scale X-ray microscopes. The article is intended to enable researchers through a high-level view of the toolbox of labels available today for studies aiming to analyze dynamic subcellular and molecular processes in cell culture systems as well as in animal tissues—and ultimately allow investigators to determine the precise location of macromolecular complexes by EM.
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Acknowledgments
The work presented here was conducted at the National Center for Microscopy and Imaging Research at San Diego, which is supported by NIH Grant GM103412 awarded to Dr. Mark Ellisman. National Institutes of Health grants P41RR004050 (Mark Ellisman) and GM086197 (Roger Y. Tsien and Mark Ellisman) and AHA Grant 10SDG2610281 (Daniela Boassa) provided funding for this research.
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Ellisman, M.H., Deerinck, T.J., Kim, K.Y. et al. Advances in molecular probe-based labeling tools and their application to multiscale multimodal correlated microscopies. J Chem Biol 8, 143–151 (2015). https://doi.org/10.1007/s12154-015-0132-6
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DOI: https://doi.org/10.1007/s12154-015-0132-6