Understanding the photophysics of the spinach-DFHBI RNA aptamer-fluorogen complex to improve live-cell RNA imaging

J Am Chem Soc. 2013 Dec 18;135(50):19033-8. doi: 10.1021/ja411060p. Epub 2013 Dec 10.


The use of aptamer-fluorogen complexes is an emerging strategy for RNA imaging. Despite its promise for cellular imaging and sensing, the low fluorescence intensity of the Spinach-DFHBI RNA aptamer-fluorogen complex hampers its utility in quantitative live-cell and high-resolution imaging applications. Here we report that illumination of the Spinach-fluorogen complex induces photoconversion and subsequently fluorogen dissociation, leading to fast fluorescence decay and fluorogen-concentration-dependent recovery. The fluorescence lifetime of Spinach-DFHBI is 4.0 ± 0.1 ns irrespective of the extent of photoconversion. We detail a low-repetition-rate illumination scheme that enables us to maximize the potential of the Spinach-DFHBI RNA imaging tag in living cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aptamers, Nucleotide / chemistry*
  • Fluorescence
  • Green Fluorescent Proteins / chemistry
  • Kinetics
  • Photochemical Processes
  • Spinacia oleracea / chemistry*


  • Aptamers, Nucleotide
  • Green Fluorescent Proteins