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New sensorless wavefront sensing approach for two photon microscopy: application to in vivo imaging of the hippocampus

Abstract : Deep in vivo two photon microscopy is severely limited by optical aberrations. Adaptive optics is now a widely used technique to overcome this issue. It relies on one of several possible wavefront sensing techniques. The modal sensorless wavefront sensing approach, initially developed for two photon imaging by Débarre et al., has become a standard technique based on the maximization of an image quality metric such as the mean image intensity. However, this technique is influenced by the sample, which may induce a strong bias on the wavefront estimation, the so-called sample dependence. This PhD work aims at developing an improved modal sensorless approach that is not affected by sample dependence. It is part of a project of in vivo neuronal imaging of the mouse hippocampus which gathers three laboratories: INMED, Institut Fresnel and Onera. I first study the impact of aberrations and of the sample structure on the mean image intensity. A new analytical expression of the mean image intensity is given and makes explicit the interplay between the shape of the 3D PSF and the sample spatial distribution. Through numerical simulations I evaluate the metric sensitivity to aberrations for different samples structures. Secondly, I study the Standard Modal Sensorless (SMS) approach for different sample scenarios. I characterize the sample dependence issue induced by strong fluorescent structures located out-of-focus. I then show that the displacement-free technique fails at fully removing the sample dependence. This analysis leads to the development of our Axially-Locked Modal Sensorless approach (ALMS). This new approach solves the sample dependence by an automatic and controlled adjustment of the focusing depth to lock on bright sample features. Furthermore, the ALMS approach is based on an image quality metric specifically designed for this axial locking. The performance of this approach is numerically compared to the SMS approaches. Finally, the ALMS approach is demonstrated through ex vivo and in vivo experimental tests in the context of neuronal imaging of the hippocampus.
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Submitted on : Thursday, January 25, 2018 - 4:47:58 PM
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Joel Teixeira. New sensorless wavefront sensing approach for two photon microscopy: application to in vivo imaging of the hippocampus. Optics [physics.optics]. Observatoire de Paris, 2017. English. ⟨tel-01693038⟩

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