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Development of a 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. Indirect or sensorless wavefront estimation has the advantage of being easy-to-implement on existing systems. Modal sensorless 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, the indirectly inferred wavefront is influenced by the sample, which may induce a strong bias in the estimation, the so-called sample dependence. This PhD work aims at developing an improved modal sensorless approach that is not affected by sample dependence.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 show that the metric sensitivity to aberrations is preserved for samples much larger than the spatial resolution.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. Then, I 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 so as to lock on bright sample features. Furthermore, the ALMS approach is based on a specifically designed image quality metric.The performance of this approach is numerically compared with the SMS and the displacement-free approaches. Finally, ALMS is demonstrated through ex vivo and in vivo experimental tests.
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Submitted on : Wednesday, October 10, 2018 - 11:00:06 AM
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Joël Teixeira. Development of a new sensorless wavefront sensing approach for two photon microscopy : application to in vivo imaging of the hippocampus. Astrophysics [astro-ph]. Université Paris sciences et lettres, 2017. English. ⟨NNT : 2017PSLEO017⟩. ⟨tel-01891917⟩

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