Biomolecule labeling and imaging with a new fluorenyl two-photon fluorescent probe

Bioconjug Chem. 2009 Oct 21;20(10):1992-2000. doi: 10.1021/bc900302t. Epub 2009 Oct 2.


Closely involved in the progression of nonlinear bioimaging is the development of optical probes for investigating biological function and activity. Introduction of new fluorescent compounds possessing enhanced nonlinearities is essential for advancing the utility of two-photon absorption (2PA) processes in the biological sciences. Herein, we report the synthesis of fluorene-based fluorophores tailored for multiphoton imaging, incorporating the succinimidyl ester and thioester functionality as reactive linkers for further coupling with a wide variety of biologically relevant molecules. The succinimidyl ester amine reactive probe was conjugated with the cyclic peptide RGDfK and polyclonal antirat IgG protein. Upon conjugation, the basic molecular architecture and photophysical properties of the active 2PA chromophore remain unchanged. Conventional and two-photon fluorescence microscopy (2PFM) imaging of COS-7 and HeLa cells, incubated with either the fluorene-RGD peptide conjugate or the fluorene-IgG conjugate, was demonstrated. The fluorene-IgG conjugate was used to image cell spindles at early mitotic developmental stages.

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

  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Fluorenes* / analysis
  • Fluorenes* / chemical synthesis
  • Fluorescent Dyes* / analysis
  • Fluorescent Dyes* / chemical synthesis
  • HeLa Cells
  • Humans
  • Imaging, Three-Dimensional
  • Immunoglobulins / chemistry*
  • Immunoglobulins / immunology
  • Microscopy, Fluorescence, Multiphoton*
  • Peptides, Cyclic / chemistry*
  • Photons


  • Fluorenes
  • Fluorescent Dyes
  • Immunoglobulins
  • Peptides, Cyclic
  • cyclic (arginyl-glycyl-aspartyl-phenylalanyl-lysyl)