Dual illuminations to reduce phototoxicity and fluorescence losses in fluorescence imaging
PhD project at ENS Paris (Department of Chemistry) in collaboration with INRAE. Start in fall 2022.
Project description:
In fluorescence imaging, the light used for excitation leads, in parallel to fluorescence, to the formation of triplet excited states of the fluorophores. These triplet states are involved as intermediates in phototoxicity and fluorophore photobleaching, two major limitations for dynamic in vivo studies. We have recently shown that it is possible to exploit a photophysical process called reverse intersystem crossing (RISC) to depopulate the triplet states of green and yellow fluorescent proteins, and thus reduce their photobleaching and the associated phototoxicity in vivo. RISC is obtained by co-illuminating the fluorophores during their excitation at a second wavelength tuned to the absorption of their triplet state.
The PhD project builds on these results and aims to further explore the potential of RISC and dual illuminations to reduce phototoxicity and fluorescence losses in fluorescence imaging. The objectives are, in particular, to extend our approach to other fluorophores, to evaluate its potential to reduce the phototoxicity due to the excitation of endogenous chromophores in live cells, and to exploit dual illuminations to control fluorophore blinking in single-molecule and super-resolution imaging.
During this project, you will acquire skills in photophysics and photochemistry, spectroscopy, kinetic modeling, live cell fluorescence imaging, expression and purification of fluorescent proteins. You will be involved in an ongoing collaboration between a team of physical chemists from ENS and a team of biologists from INRAE.
Required background:
Candidates should hold a Master’s degree in Physical Chemistry or Biophysics and have a strong interest in the interface with Biology. Previous experience in photophysics, photochemistry, spectroscopy, kinetics, fluorescence imaging and/or the handling of live biological samples will be a plus.
Financial support:
Financial support is expected from a grant from ED 388 and will be confirmed by the end of April. We have no local candidate and are very well placed at the laboratory level to obtain this grant.
References:
- A. Espagne, K. Brettel, L. Jullien, T. Le Saux, L. Ludvikova, P. Müller & L. Robert, Method for enhancing the photoresistance of a fluorescent species and fluorescence microscopy system suitable for implementing said method. Patent application N° EP21305834.0 (2021).
- L. Ludvikova, T. Panier, M.-A. Plamont, A. Tebo, L. Jullien, L. Robert, T. Le Saux & A. Espagne, Near-infrared co-illumination of fluorescent proteins reduces photobleaching and phototoxicity, under review at Nat. Biotechnol.
Contacts:
ENS : A. Espagne (agathe.espagne@ens.psl.eu) & T. Le Saux (thomas.lesaux@ens.psl.eu)
INRAE : L. Robert (lydia.robert@inrae.fr)