A150 - Adaptive optics two-photon fluorescence microscopy for in depth bioimaging

Alice Guillaume-manca (alice.guillaume-manca@espci.fr)
Mathias Mercier (mathias.mercier@espci.fr)

This workshop aims to explain and demonstrate the gain of adaptive optics fluorescence microscopy for biological samples imaging. Indeed, image quality deep inside a sample is degraded by the inhomogeneities of the biological tissues which strongly distort the phase of optical waves, i.e. the wavefront of the light of interest, and thus limit contrast, spatial resolution and sensitivity. In the recent years, adaptive optics, which has been initially developed for astronomy, has shown its ability to significantly increase signal and resolution deep inside biological tissues, by correcting the optical aberrations induced by the sample itself. This is achieved through 1) wavefront sensing and 2) correction using optical wavefront modulators like deformable mirrors. Two main adaptive optics approaches have been proposed in microscopy, based on two methods of wavefront estimation before correction. This workshop will present the physical concepts underlying adaptive optics with simple experiments and explain the existing strategies to implement it in microscopy, with their benefits and constraints. We will then focus on an example of an adaptive optics loop implementation in two photon microscopy based on a novel wavefront measurement approach, with an extended source Shack-Hartmann sensor. This original approach leads to an efficient and fast correction in scattering samples and allows to correct aberrations up to several hundred microns deep inside a fixed mouse brain tissue, proving large gain in intensity and resolution. At the end of this workshop, the participants will be able to evaluate if adaptive optics can bring image enhancement regarding the microscopy technique they use and their bioimaging application.