Dynamic full-field optical coherence tomography: 3D live-imaging of retinal organoids

Optical coherence tomography offers astounding opportunities to image the complex structure of living tissue, but lacks functional information. We present dynamic full-field optical coherence tomography to image living human induced pluripotent stem cell-derived retinal organoids non-invasively. Colored images with an endogenous contrast linked to organelle motility are generated, with sub-micrometer spatial resolution and millisecond temporal resolution, opening an avenue to identify specific cell types in living tissue via their function.Comment: 14 pages, 5 figures, 1 table, 6 video

Multimodal 3D optical tomography : spatio-temporal processings, wavefront correction and automatic classification.

Cette thèse vise à appliquer et combiner des méthodes numériques et optiques pour repousser les limites de la tomographie optique cohérente plein champ (FFOCT) statique et dynamique pour la microscopie et l'imagerie médicale. Des méthodes de post-traitement utilisant la décomposition en valeurs singulières ont permis pour la première fois l'acquisition d'images dynamiques in vivo tandis que l'utilisation des signaux non stationnaires a permis d'obtenir des images avec un meilleur rapport signal sur bruit, soit la possibilité d'imager plus profondément les échantillons. L'application de l'imagerie dynamique est présentée sur des organoïdes rétiniens où nous montrons que notre méthode est capable de fournir de nouvelles perceptions biologiques intéressantes qui ne sont possibles avec aucune autre méthode. Des développements matériel pour compenser les aberrations optiques ont été menés avec succès, ce qui a permis une mise en œuvre peu complexe et peu coûteuse permettant d'acquérir de manière fiable des images de la rétine avec une résolution en limite de diffraction. La compréhension de la manifestation des aberrations optiques en FFOCT validée expérimentalement nous a permis de concevoir et de simuler les performances du système proposé. Enfin, les applications cliniques potentielles du FFOCT dynamique et statique pour l'angiographie de l'œil humain in vivo, la cicatrisation ex vivo, la classification des cellules de la rétine et le dépistage du cancer du sein par des méthodes d'apprentissage automatique ont été démontrées avec succès.This PhD project aims at combining numerical and optical methods to apply and push the limits of static and dynamic full-field optical coherent tomography (FFOCT) for microscopy and medical imaging. Post-processing methods using singular value decomposition allowed the acquisition of dynamic images in vivo for the first time while the use of the signals non-stationarities allowed to image with a better signal to noise ratio, hence deeper inside samples. Application of dynamic imaging is presented on retinal organoids where we show that our method is able to provide new interesting biological insights that are not possible with any other methods. Hardware developments to counteracts optical aberrations were successfully conducted leading to low complexity and cost efficient implementation which can reliably acquire retinal images with a diffraction limited resolution. The understanding and demonstration of the particular aberrations manifestation in FFOCT allowed us to design and simulate the performances of the proposed system. Finally, potential clinical applications of dynamic and static FFOCT for angiography in the human eye in vivo, wound healing ex vivo, retinal cell classification and breast cancer screening using machine learning methods are successfully demonstrated

Imagerie optique 3D multimodale : traitements spatio-temporels, correction du front d'onde et classification automatique.

This PhD project aims at combining numerical and optical methods to apply and push the limits of static and dynamic full-field optical coherent tomography (FFOCT) for microscopy and medical imaging. Post-processing methods using singular value decomposition allowed the acquisition of dynamic images in vivo for the first time while the use of the signals non-stationarities allowed to image with a better signal to noise ratio, hence deeper inside samples. Application of dynamic imaging is presented on retinal organoids where we show that our method is able to provide new interesting biological insights that are not possible with any other methods. Hardware developments to counteracts optical aberrations were successfully conducted leading to low complexity and cost efficient implementation which can reliably acquire retinal images with a diffraction limited resolution. The understanding and demonstration of the particular aberrations manifestation in FFOCT allowed us to design and simulate the performances of the proposed system. Finally, potential clinical applications of dynamic and static FFOCT for angiography in the human eye in vivo, wound healing ex vivo, retinal cell classification and breast cancer screening using machine learning methods are successfully demonstrated.Cette thèse vise à appliquer et combiner des méthodes numériques et optiques pour repousser les limites de la tomographie optique cohérente plein champ (FFOCT) statique et dynamique pour la microscopie et l'imagerie médicale. Des méthodes de post-traitement utilisant la décomposition en valeurs singulières ont permis pour la première fois l'acquisition d'images dynamiques in vivo tandis que l'utilisation des signaux non stationnaires a permis d'obtenir des images avec un meilleur rapport signal sur bruit, soit la possibilité d'imager plus profondément les échantillons. L'application de l'imagerie dynamique est présentée sur des organoïdes rétiniens où nous montrons que notre méthode est capable de fournir de nouvelles perceptions biologiques intéressantes qui ne sont possibles avec aucune autre méthode. Des développements matériel pour compenser les aberrations optiques ont été menés avec succès, ce qui a permis une mise en œuvre peu complexe et peu coûteuse permettant d'acquérir de manière fiable des images de la rétine avec une résolution en limite de diffraction. La compréhension de la manifestation des aberrations optiques en FFOCT validée expérimentalement nous a permis de concevoir et de simuler les performances du système proposé. Enfin, les applications cliniques potentielles du FFOCT dynamique et statique pour l'angiographie de l'œil humain in vivo, la cicatrisation ex vivo, la classification des cellules de la rétine et le dépistage du cancer du sein par des méthodes d'apprentissage automatique ont été démontrées avec succès

Adaptive-glasses time-domain FFOCT for wide-field high-resolution retinal imaging with increased SNR

International audienceThe highest three-dimensional (3D) resolution possible in in vivo retinal imaging is achieved by combining optical coherence tomography (OCT) and adaptive optics. However, this combination brings important limitations, such as small field-of-view and complex, cumbersome systems, preventing so far the translation of this technology from the research lab to clinics. In this Letter, we mitigate these limitations by combining our compact time-domain full-field OCT (FFOCT) with a multi-actuator adaptive lens positioned just in front of the eye, in a technique we call the adaptive-glasses wavefront sensorless approach. Through this approach, we demonstrate that ocular aberrations can be corrected, increasing the FFOCT signal-to-noise ratio (SNR) and enabling imaging of different retinal layers with a 3D cellular resolution over a 5∘×5∘ field-of-view, without apparent anisoplanatism

Coherence gate shaping for wide field high-resolution in vivo retinal imaging with full-field OCT

International audienceAllying high-resolution with a large field-of-view (FOV) is of great importance in the fields of biology and medicine, but it is particularly challenging when imaging non-flat living samples such as the human retina. Indeed, high-resolution is normally achieved with adaptive optics (AO) and scanning methods, which considerably reduce the useful FOV and increase the system complexity. An alternative technique is time-domain full-field optical coherence tomography (FF-OCT), which has already shown its potential for in-vivo high-resolution retinal imaging. Here, we introduce coherence gate shaping for FF-OCT, to optically shape the coherence gate geometry to match the sample curvature, thus achieving a larger FOV than previously possible. Using this instrument, we obtained high-resolution images of living human photoreceptors close to the foveal center without AO and with a 1 mm × 1 mm FOV in a single shot. This novel advance enables the extraction of photoreceptor-based biomarkers with ease and spatiotemporal monitoring of individual photoreceptors. We compare our findings with AO-assisted ophthalmoscopes, highlighting the potential of FF-OCT, as a compact system, to become a routine clinical imaging technique

Real-time non-contact cellular imaging and angiography of human cornea and limbus with common-path full-field/SD OCT

International audienceIn today's clinics, a cell-resolution view of the cornea can be achieved only with a confocal microscope (IVCM) in contact with the eye. Here, we present a common-path full-field/spectral-domain OCT microscope (FF/SD OCT), which enables cell-detail imaging of the entire ocular surface in humans (central and peripheral cornea, limbus, sclera, tear film) without contact and in real-time. Real-time performance is achieved through rapid axial eye tracking and simultaneous defocusing correction. Images contain cells and nerves, which can be quantified over a millimetric field-of-view, beyond the capability of IVCM and conventional OCT. In the limbus, palisades of Vogt, vessels, and blood flow can be resolved with high contrast without contrast agent injection. The fast imaging speed of 275 frames/s (0.6 billion pixels/s) allows direct monitoring of blood flow dynamics, enabling creation of high-resolution velocity maps. Tear flow velocity and evaporation time can be measured without fluorescein administration

Dynamic full-field optical coherence tomography: 3D live-imaging of retinal organoids

International audienceAbstract Optical coherence tomography offers astounding opportunities to image the complex structure of living tissue but lacks functional information. We present dynamic full-field optical coherence tomography as a technique to noninvasively image living human induced pluripotent stem cell-derived retinal organoids. Coloured images with an endogenous contrast linked to organelle motility are generated, with submicrometre spatial resolution and millisecond temporal resolution, creating a way to identify specific cell types in living tissue via their function