Dual-color deep-tissue three-photon microscopy with a multiband infrared laser

Khmaies Guesmi; Lamiae Abdeladim; Samuel Tozer; Pierre Mahou; Takuma Kumamoto; Karolis Jurkus; Philippe Rigaud; Karine Loulier; Nicolas Dray; Patrick Georges; Marc Hanna; Jean Livet; Willy Supatto; Emmanuel Beaurepaire; Frédéric Druon

doi:10.1038/s41377-018-0012-2

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Dual-color deep-tissue three-photon microscopy with a multiband infrared laser

Original Articles

- Dual-color deep-tissue three-photon microscopy with a multiband infrared laser
- Light: Science & Applications Vol. 7, Issue 4, Pages: 119-127(2018)
- Affiliations：
  
  1.Laboratory Charles Fabry, Institut d’Optique Graduate School, CNRS, Université Paris-Saclay, 91128 Palaiseau, France
  2.Laboratory for Optics and Biosciences, Ecole Polytechnique, CNRS, INSERM, 91128 Palaiseau, France
  3.Vision Institute, CNRS Sorbonne Universités, Université Paris 6, INSERM, 75012 Paris, France
  4.Zebrafish Neurogenetics Unit, Developmental and Stem Cell Biology Department, Institut Pasteur, CNRS, 75015 Paris, France
- Author bio：
  
  Emmanuel Beaurepaire (emmanuel.beaurepaire@polytechnique.edu)
  Frédéric. Druon (frederic.druon@institutoptique.fr)
- Funds：
- DOI：10.1038/s41377-018-0012-2
  CLC：
- Published：2018，
  
  Published Online：06 June 2018，
  
  Received：20 September 2017，
  
  Revised：12 February 2018，
  
  Accepted：23 February 2018
- Accepted：
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Guesmi, K. et al. Dual-color deep-tissue three-photon microscopy with a multiband infrared laser. Light: Science & Applications, 4, 119-127 (2018).
DOI：

Guesmi, K. et al. Dual-color deep-tissue three-photon microscopy with a multiband infrared laser. Light: Science & Applications, 4, 119-127 (2018). DOI： 10.1038/s41377-018-0012-2.

摘要

Abstract

Multiphoton microscopy combined with genetically encoded fluorescent indicators is a central tool in biology. Three-photon (3P) microscopy with excitation in the short-wavelength infrared (SWIR) water transparency bands at 1.3 and 1.7 µm opens up new opportunities for deep-tissue imaging. However

novel strategies are needed to enable in-depth multicolor fluorescence imaging and fully develop such an imaging approach. Here

we report on a novel multiband SWIR source that simultaneously emits ultrashort pulses at 1.3 and 1.7 µm that has characteristics optimized for 3P microscopy: sub-70 fs duration

1.25 MHz repetition rate

and µJ-range pulse energy. In turn

we achieve simultaneous 3P excitation of green fluorescent protein (GFP) and red fluorescent proteins (mRFP

mCherry

tdTomato) along with third-harmonic generation. We demonstrate in-depth dual-color 3P imaging in a fixed mouse brain

chick embryo spinal cord

and live adult zebrafish brain

with an improved signal-to-background ratio compared to multicolor two-photon imaging. This development opens the way towards multiparametric imaging deep within scattering tissues.

关键词

Keywords

references

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