© THOMAS DEERINCK, NCMIR/SCIENCE SOURCEFluorescent dyes and genetic tags have revolutionized researchers’ ability to determine the location of specific proteins in space and time using light microscopy (LM). With the advent of super-resolution fluorescence LM techniques, much more precise location of proteins within a cell is possible, but those techniques do not provide information about the ultrastructural cellular context of biomolecules. Electron microscopy (EM) is an unbeatable tool for providing structural data at the nanometer scale, but the field of view in EM is very narrow and the molecular scenery is highly crowded, making it tedious and time-consuming to locate specific structures and cellular events, especially if they are rare. Moreover, EM can only provide a static snapshot of the sample because cells and tissues must be fixed prior to imaging.
Recently, researchers have developed methods that allow both light- and electron-microscopic examination of the same specimen, an approach known as correlated light and electron microscopy, or CLEM. LM imaging (which is typically performed before EM because the electron beam usually wipes out fluorescent signals) provides an overview of the sample, enabling researchers to identify structures and regions of interest for investigation with EM.
One of the primary challenges in CLEM, however, is locating the same area that was imaged using LM, says Chris Arthur, a senior research scientist at FEI Company in Hillsboro, Oregon, which manufacturers several different tools for CLEM. Even as researchers manage to home in on the region of interest, it can be difficult to accurately overlay the two types of images, ...
