Multicolor Super-Resolution Imaging with Photo-Switchable Fluorescent Probes
Mark Bates,1
Bo Huang,2,3
Graham T. Dempsey,4
Xiaowei Zhuang2,3,5*
Recent advances in far-field optical nanoscopy have enabled fluorescence imaging with a spatial resolution of 20 to 50 nanometers. Multicolor super-resolution imaging, however, remains a challenging task. Here, we introduce a family of photo-switchable fluorescent probes and demonstrate multicolor stochastic optical reconstruction microscopy (STORM). Each probe consists of a photo-switchable "reporter" fluorophore that can be cycled between fluorescent and dark states, and an "activator" that facilitates photo-activation of the reporter. Combinatorial pairing of reporters and activators allows the creation of probes with many distinct colors. Iterative, color-specific activation of sparse subsets of these probes allows their localization with nanometer accuracy, enabling the construction of a super-resolution STORM image. Using this approach, we demonstrate multicolor imaging of DNA model samples and mammalian cells with 20- to 30-nanometer resolution. This technique will facilitate direct visualization of molecular interactions at the nanometer scale.
1 School of Engineering and Applied Sciences, Harvard University, Cambridge, MA 02138, USA.
2 Howard Hughes Medical Institute, Harvard University, Cambridge, MA 02138, USA.
3 Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138, USA.
4 Graduate Program in Biophysics, Harvard University, Cambridge, MA 02138, USA.
5 Department of Physics, Harvard University, Cambridge, MA 02138, USA.
* To whom correspondence should be addressed. E-mail: zhuang{at}chemistry.harvard.edu
