In the conventional electron micrograph at left, it is difficult to distinguish form and function of two hippocampal astrocytes in a mouse brain. The multicolor micrograph at right marks the two astrocytes using different color lanthanides and more clearly shows where they contact and nurture the same synapse (dark railroad lines). Credit: Image courtesy of Stephen Adams, NCMIR, UC San Diego
Electron microscopy (EM), which uses particle beams of accelerated electrons to interrogate specimens, has long been a leading technology for revealing the shape and structure of the tiniest objects, from the cells which make up the bodies organs and microbes to individual building blocks or molecules which comprise cells, in often dramatic 3D detail. But current EM techniques are limited in that they produce images only in grayscale, with colorization added later. Researchers at University of California San Diego School of Medicine and Howard Hughes Medical Institute describe a new form of multicolor EM that allows for simultaneous visualization of multiple molecular species.
“It reveals details, actions and processes that aren’t necessarily visible – or even suspected – in a more monochromatic view,” said Prof. Mark H. Ellisman, PhD. Although there have been major improvements in multicolor and super-resolution fluorescence microscopy, progress in EM has been limited, achieved through automation and developments eg miniSOG protein, a new type of genetic tag visible under an EM microscope developed by Tsien, Ellisman and colleagues in 2011.
To create multicolor EM images, researchers sequentially painted cellular structures such as proteins, membranes or whole cells with different “rare earth” metals, such as lanthanum, cerium and praseodymium in the form of precipitates. “A transmission electron microscope can distinguish each of these metals by electron energy-loss to give elemental maps of each that can be overlaid in color on the familiar monochrome electron micrograph,” said Adams. “Each color highlights a different component of the cellular ultrastructure.”
Multicolor EM offers the possibility to differentiate detail not possible with standard EM, which uses gold particles to label structures but which appear in images as sometimes hard-to-distinguish black spots. It provides spatial resolution not possible with fluorescence microscopy. Adams said: “In theory, we should be able to add many more colors if we can develop more ways of precipitating additional lanthanides. The method is quite simple to do, uses easily made chemicals and requires detectors that are already present on many transmission electron microscopes so it is potentially readily transferable to other laboratories. Further research is needed to improve the chemistry and sensitivity of the method, but this work will hopefully inspire other groups to devise similar methods in this field.” https://health.ucsd.edu/news/releases/Pages/2016-11-03-new-technique-visualizes-mulitple-objects-in-many-colors.aspx
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