Dye Colors Up Live Cell Surfaces In 3-D

Many organic dyes otherwise suitable for superresolution microscopy require ultraviolet light to turn on their fluorescence, which can damage live cells. A new rhodamine spirolactam, developed by W. E. Moerner of Stanford University, Robert J. Twieg of Kent State University, and coworkers, is more compatible with live-cell imaging because it switches from its nonfluorescent form to its fluorescent isomer when irradiated with visible light. The researchers have used the dye to acquire three-dimensional images of commonly studied Caulobacter crescentus bacteria cells (J. Am. Chem. Soc. 2014, DOI: 10.1021/ja508028h). The positively charged dye, which doesn’t cross the bacterial cell membrane, contains a N-hydroxysuccinimide ester that covalently labels amines on the cell surface. The researchers turn on the fluorescence of a few dye molecules at a time with a low-intensity 405-nm purple laser and read the fluorescence with a 561-nm green laser. By doing this many times, they build superresolution images of the live cell surfaces in which they are able to resolve stalk structures that aren’t visible using standard microscopy.