Master Projects (M1 or M2)

Single Molecule Localization Microscopy (SMLM) is a powerful method to achieve nanometer resolution in biological samples using a simple widefield optical microscope. It relies on inducing stochastic blinking in the fluorophores used to label the structure of interest by using chemical buffers that affect their photophysics. Our aim is to develop methods to improve super-resolved tissue imaging by focusing on the buffer composition. We recently made some good progress on mutilcolor imaging [1,2], and now aim at improving the achievable resolution for some fluorophores by further tuning this buffer, using both ensemble and single molecule experiments to extract photophysical parameters.

Second harmonic generation (SHG) microscopy enables imaging of collagen without any labelling and with unequalled sensitivity in intact tissues. This is directly related to the property of SHG to be nonzero only in dense and non-centrosymmetrical materials. Nevertheless, the build-up of SHG is a complex issue because of its coherent nonlinear nature and of the heterogeneity of collagen distribution in tissues. Collagen is indeed organized as fibrils, which size and 3D organization is specific to each tissue (skin, artery, lung, bone…). Extracting as much information as possible about the structure of a tissue from SHG images is therefore an active research field. An effective method consists in varying the excitation polarization, which yields the collagen orientation in every pixel. Nevertheless, numerical simulations are needed to model this polarimetric SHG (pSHG) signal for realistic tissue geometries and reconstruct in a reliable way the collagen disctribution at sub-micrometer scale. This project aims at modeling pSHG in the Human cornea and compare these numerical simulations to pSHG images recorded recently by our group. Two approaches will be implemented, based on previous work at LOB [3,4]:

(i) a semi-analytical approach based on Green’s functions, which considers only nonlinear indices;

(ii) a full numerical approach using Finite Difference Time-Domain (FDTD) to take into account nanometer-scale heterogeneities in the refractive index that may affect the wave propagation.

The internship mainly aims at developing Green Function and FDTD pipelines for simulating the anisotropy of collagen fibrils and the heterogeneity of their 3D distribution.

Nonlinear microscopy is mostly used for live tissue imaging, and resolution is usually not the predominant preoccupation. However, it is sometimes useful to have the ability to zoom in on a particular biological process, hence the need to develop super-resolution approaches. The aim of this project is to perform numerical simulation for different nonlinear processes involving 2 different shaped beams, which should provide enough degrees of freedom to at least double the resolution (or to provide isotropic resolution).

When looking at sparse objects, or when quantifying the overall concentration of a protein, it is sometimes useful to degrade the axial resolution of a microscope to image a larger volume at once. Achieving this without degrading the lateral resolution is possible, and we will consider different experimental approach that rely on the coherence of the excitation and detected light, first in the case of super-resolution microscopy and if time allows for nonlinear microscopy.

PhD Project

Some of these projects described above could provide the basis of a PhD projects, contact me for more details. I have no funding available on my side, so the candidates would be expected to apply to the IPP graduate school fellowships. PhD Track students are welcome: www.ip-paris.fr/education/phd-track/phd-track-physique

Post-doc

I sadly do not have any funding currently for post-doc positions, but anyone wishing to apply for postdoctoral fellowship related to the group’s interests is welcome to contact me

References:

1 - An Optimized Buffer for Repeatable Multicolor STORM Vaky Abdelsayed, Hadjer Boukhatem, & Nicolas Olivier (2022). Now also peer-reviewed (paywalled)

2 - Evaluation of Slowfade Diamond as a buffer for STORM microscopy Hadjer Boukhatem, Beatrice Durel, Manon Raimbault, Audrey Laurent, and Nicolas Olivier (2023)

3 - Harmonic microscopy of isotropic and anisotropic microstructure of the human cornea N. Olivier, et al. (2010)

4 - Modeling nonlinear microscopy near index-mismatched interfaces J. Morizet, G. Sartorello, N. Dray, C. Stringari, E. Beaurepaire, N. Olivier (2021)