Monomeric red fluorescent proteins with a large Stokes shift

Kiryl D. Piatkevich, James Hulit, Oksana M. Subach, Bin Wu, Arian Abdulla, Jeffrey E. Segall, Vladislav Verkhusha

Research output: Contribution to journalArticle

103 Citations (Scopus)

Abstract

Two-photon microscopy has advanced fluorescence imaging of cellular processes in living animals. Fluorescent proteins in the blue-green wavelength range are widely used in two-photon microscopy; however, the use of red fluorescent proteins is limited by the low power output of Ti-Sapphire lasers above 1,000 nm. To overcome this limitation we have developed two red fluorescent proteins, LSS-mKate1 and LSS-mKate2, which possess large Stokes shifts with excitation/emission maxima at 463/624 and 460/605 nm, respectively. These LSS-mKates are characterized by high pH stability, photostability, rapid chromophore maturation, and monomeric behavior. They lack absorbance in the green region, providing an additional red color to the commonly used red fluorescent proteins. Substantial overlap between the two-photon excitation spectra of the LSS-mKates and blue-green fluorophores enables multicolor imaging using a single laser. We applied this approach to a mouse xenograft model of breast cancer to intravitally study the motility and Golgi-nucleus alignment of tumor cells as a function of their distance from blood vessels. Our data indicate that within 40 μm the breast cancer cells show significant polarization towards vessels in living mice.

Original languageEnglish (US)
Pages (from-to)5369-5374
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume107
Issue number12
DOIs
StatePublished - Mar 23 2010

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Photons
Microscopy
Lasers
Breast Neoplasms
Aluminum Oxide
Optical Imaging
Heterografts
Blood Vessels
Color
red fluorescent protein
Neoplasms
Proteins

Keywords

  • Cell polarity
  • Intravital imaging
  • Keima
  • mKate
  • Two-photon microscopy

ASJC Scopus subject areas

  • General

Cite this

Monomeric red fluorescent proteins with a large Stokes shift. / Piatkevich, Kiryl D.; Hulit, James; Subach, Oksana M.; Wu, Bin; Abdulla, Arian; Segall, Jeffrey E.; Verkhusha, Vladislav.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 107, No. 12, 23.03.2010, p. 5369-5374.

Research output: Contribution to journalArticle

Piatkevich, Kiryl D. ; Hulit, James ; Subach, Oksana M. ; Wu, Bin ; Abdulla, Arian ; Segall, Jeffrey E. ; Verkhusha, Vladislav. / Monomeric red fluorescent proteins with a large Stokes shift. In: Proceedings of the National Academy of Sciences of the United States of America. 2010 ; Vol. 107, No. 12. pp. 5369-5374.
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