12 research outputs found

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

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    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

    Imagerie de la rétine : suivi de structure et de l'activité cellulaire par OCT Plein Champ statique et dynamique

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    This thesis aims to demonstrate the usefulness of static and dynamic Full-Field Optical Coherence Tomography (FFOCT) for retinal imaging, with in vitro (cell cultures in 2D or 3D), ex vivo (retinas dissected from different species) and in vivo (direct imaging of the eye of the patient) samples.By stabilising the three dimensional and timelapse dynamic acquisitions, it allows a better following and interpretation of the evolution of in vitro samples, such as retinal organoids (produced from human pluripotent stem cells) or retinal pigment epithelium cell cultures (from pluripotent stem cells and primary porcine cells). The recorded dynamic signals can be differentiated by the cell type (photoreceptors, retinal pigment epithelium, etc.) or the expressed phenotypes (for example dying cell, cell division, etc.), allowing a totally non invasive imaging in biology. These signals, representing the cellular activity, come from organelles present in the cells, especially mitochondria, and help to an easy interpretation of the condition of the cells.This innovative contrast allows to study retinal degenerative diseases in vitro, such as Age-related Macular Degeneration or Retinitis Pigmentosa. Disease modeling can easily be studied by evaluating the cell response over time, for example after an induced stress on the cell culture (scratch assays). Drug screening for these diseases will also be eased by following the evolution of the dynamic signal produced by the cells after treatment.Finally, the application of Full-Field Optical Coherence Tomography to in vivo retinal imaging permit a non invasive study of the different layers of the retina (from the nerves to the retinal pigment epithelium). It is a technique less complex than those with adaptive optics usually used for in vivo imaging. The stabilisation and signal improvement justify the idea of the future in vivo implementation of dynamic FFOCT for the detection of diseases at early stages, such as Age-related Macular Degeneration or Glaucoma.Cette th√®se cherche √† d√©montrer l'utilit√© de la Tomographie en Coh√©rence Optique (OCT) plein champ statique et dynamique pour l'imagerie de la r√©tine, qu'il s'agisse d'√©chantillons in vitro (cultures cellulaires 2D ou 3D), ex vivo (r√©tines extraites de diff√©rentes esp√®ces) ou in vivo (imagerie direct de l'Ňďil du patient). La stabilisation des acquisitions dynamiques, √† la fois 3D et temporelle, permet un meilleur suivi et une meilleure interpr√©tation de l'√©volution d'√©chantillons in vitro, tels que les organo√Įdes de r√©tine (produits √† partir de cellules souches pluripotentes humaines) ou les cultures cellulaires d'√©pith√©lium pigmentaire r√©tinien (√† partir de cellules souches humaines et de cellules primaires de porc). Les signaux dynamiques obtenus diff√®rent selon le type cellulaire (photor√©cepteurs, √©pith√©lium pigmentaire r√©tinien, etc.), mais √©galement selon les ph√©notypes exprim√©s (mort cellulaire, division cellulaire, etc.), permettant une imagerie totalement non invasive en biologie. Ces signaux, repr√©sentant l'activit√© cellulaire, proviennent des organelles pr√©sentes dans les cellules, et notamment des mitochondries, et aide √† l'interpr√©tation de l'√©tat des cellules.Ce contraste innovant permettra d'√©tudier des maladies d√©g√©n√©ratives de la r√©tine in vitro, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou la R√©tinite Pigmentaire. La mod√©lisation de maladies peut √™tre facilement √©tudi√© en √©valuant la r√©ponse des cellules au cours du temps, par exemple apr√®s un stress induit √† la culture cellulaire. La recherche de traitements ad√©quats pour ces maladies sera aussi facilit√©e par le suivi de l'√©volution du signal dynamique des cellules, apr√®s traitement.Enfin l'application de l'OCT plein champ √† l'imagerie in vivo de la r√©tine permet une √©tude non invasive des diff√©rentes couches de la r√©tine (des nerfs jusqu'√† l'√©pith√©lium pigmentaire r√©tinien). Elle est aussi moins complexe que les techniques d'optique adaptative habituellement utilis√©es. Les am√©liorations de stabilisation et de signal justifient l'id√©e d'impl√©menter l'OCT plein champ dynamique in vivo dans le futur pour une d√©tection pr√©coce des maladies, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou le Glaucome

    Imagerie de la rétine : suivi de structure et de l'activité cellulaire par OCT Plein Champ statique et dynamique

    No full text
    This thesis aims to demonstrate the usefulness of static and dynamic Full-Field Optical Coherence Tomography (FFOCT) for retinal imaging, with in vitro (cell cultures in 2D or 3D), ex vivo (retinas dissected from different species) and in vivo (direct imaging of the eye of the patient) samples.By stabilising the three dimensional and timelapse dynamic acquisitions, it allows a better following and interpretation of the evolution of in vitro samples, such as retinal organoids (produced from human pluripotent stem cells) or retinal pigment epithelium cell cultures (from pluripotent stem cells and primary porcine cells). The recorded dynamic signals can be differentiated by the cell type (photoreceptors, retinal pigment epithelium, etc.) or the expressed phenotypes (for example dying cell, cell division, etc.), allowing a totally non invasive imaging in biology. These signals, representing the cellular activity, come from organelles present in the cells, especially mitochondria, and help to an easy interpretation of the condition of the cells.This innovative contrast allows to study retinal degenerative diseases in vitro, such as Age-related Macular Degeneration or Retinitis Pigmentosa. Disease modeling can easily be studied by evaluating the cell response over time, for example after an induced stress on the cell culture (scratch assays). Drug screening for these diseases will also be eased by following the evolution of the dynamic signal produced by the cells after treatment.Finally, the application of Full-Field Optical Coherence Tomography to in vivo retinal imaging permit a non invasive study of the different layers of the retina (from the nerves to the retinal pigment epithelium). It is a technique less complex than those with adaptive optics usually used for in vivo imaging. The stabilisation and signal improvement justify the idea of the future in vivo implementation of dynamic FFOCT for the detection of diseases at early stages, such as Age-related Macular Degeneration or Glaucoma.Cette th√®se cherche √† d√©montrer l'utilit√© de la Tomographie en Coh√©rence Optique (OCT) plein champ statique et dynamique pour l'imagerie de la r√©tine, qu'il s'agisse d'√©chantillons in vitro (cultures cellulaires 2D ou 3D), ex vivo (r√©tines extraites de diff√©rentes esp√®ces) ou in vivo (imagerie direct de l'Ňďil du patient). La stabilisation des acquisitions dynamiques, √† la fois 3D et temporelle, permet un meilleur suivi et une meilleure interpr√©tation de l'√©volution d'√©chantillons in vitro, tels que les organo√Įdes de r√©tine (produits √† partir de cellules souches pluripotentes humaines) ou les cultures cellulaires d'√©pith√©lium pigmentaire r√©tinien (√† partir de cellules souches humaines et de cellules primaires de porc). Les signaux dynamiques obtenus diff√®rent selon le type cellulaire (photor√©cepteurs, √©pith√©lium pigmentaire r√©tinien, etc.), mais √©galement selon les ph√©notypes exprim√©s (mort cellulaire, division cellulaire, etc.), permettant une imagerie totalement non invasive en biologie. Ces signaux, repr√©sentant l'activit√© cellulaire, proviennent des organelles pr√©sentes dans les cellules, et notamment des mitochondries, et aide √† l'interpr√©tation de l'√©tat des cellules.Ce contraste innovant permettra d'√©tudier des maladies d√©g√©n√©ratives de la r√©tine in vitro, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou la R√©tinite Pigmentaire. La mod√©lisation de maladies peut √™tre facilement √©tudi√© en √©valuant la r√©ponse des cellules au cours du temps, par exemple apr√®s un stress induit √† la culture cellulaire. La recherche de traitements ad√©quats pour ces maladies sera aussi facilit√©e par le suivi de l'√©volution du signal dynamique des cellules, apr√®s traitement.Enfin l'application de l'OCT plein champ √† l'imagerie in vivo de la r√©tine permet une √©tude non invasive des diff√©rentes couches de la r√©tine (des nerfs jusqu'√† l'√©pith√©lium pigmentaire r√©tinien). Elle est aussi moins complexe que les techniques d'optique adaptative habituellement utilis√©es. Les am√©liorations de stabilisation et de signal justifient l'id√©e d'impl√©menter l'OCT plein champ dynamique in vivo dans le futur pour une d√©tection pr√©coce des maladies, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou le Glaucome

    Imagerie de la rétine : suivi de structure et de l'activité cellulaire par OCT Plein Champ statique et dynamique

    No full text
    Cette th√®se cherche √† d√©montrer l'utilit√© de la Tomographie en Coh√©rence Optique (OCT) plein champ statique et dynamique pour l'imagerie de la r√©tine, qu'il s'agisse d'√©chantillons in vitro (cultures cellulaires 2D ou 3D), ex vivo (r√©tines extraites de diff√©rentes esp√®ces) ou in vivo (imagerie direct de l'Ňďil du patient). La stabilisation des acquisitions dynamiques, √† la fois 3D et temporelle, permet un meilleur suivi et une meilleure interpr√©tation de l'√©volution d'√©chantillons in vitro, tels que les organo√Įdes de r√©tine (produits √† partir de cellules souches pluripotentes humaines) ou les cultures cellulaires d'√©pith√©lium pigmentaire r√©tinien (√† partir de cellules souches humaines et de cellules primaires de porc). Les signaux dynamiques obtenus diff√®rent selon le type cellulaire (photor√©cepteurs, √©pith√©lium pigmentaire r√©tinien, etc.), mais √©galement selon les ph√©notypes exprim√©s (mort cellulaire, division cellulaire, etc.), permettant une imagerie totalement non invasive en biologie. Ces signaux, repr√©sentant l'activit√© cellulaire, proviennent des organelles pr√©sentes dans les cellules, et notamment des mitochondries, et aide √† l'interpr√©tation de l'√©tat des cellules.Ce contraste innovant permettra d'√©tudier des maladies d√©g√©n√©ratives de la r√©tine in vitro, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou la R√©tinite Pigmentaire. La mod√©lisation de maladies peut √™tre facilement √©tudi√© en √©valuant la r√©ponse des cellules au cours du temps, par exemple apr√®s un stress induit √† la culture cellulaire. La recherche de traitements ad√©quats pour ces maladies sera aussi facilit√©e par le suivi de l'√©volution du signal dynamique des cellules, apr√®s traitement.Enfin l'application de l'OCT plein champ √† l'imagerie in vivo de la r√©tine permet une √©tude non invasive des diff√©rentes couches de la r√©tine (des nerfs jusqu'√† l'√©pith√©lium pigmentaire r√©tinien). Elle est aussi moins complexe que les techniques d'optique adaptative habituellement utilis√©es. Les am√©liorations de stabilisation et de signal justifient l'id√©e d'impl√©menter l'OCT plein champ dynamique in vivo dans le futur pour une d√©tection pr√©coce des maladies, telles que la D√©g√©n√©rescence Maculaire Li√©e √† l'Age ou le Glaucome.This thesis aims to demonstrate the usefulness of static and dynamic Full-Field Optical Coherence Tomography (FFOCT) for retinal imaging, with in vitro (cell cultures in 2D or 3D), ex vivo (retinas dissected from different species) and in vivo (direct imaging of the eye of the patient) samples.By stabilising the three dimensional and timelapse dynamic acquisitions, it allows a better following and interpretation of the evolution of in vitro samples, such as retinal organoids (produced from human pluripotent stem cells) or retinal pigment epithelium cell cultures (from pluripotent stem cells and primary porcine cells). The recorded dynamic signals can be differentiated by the cell type (photoreceptors, retinal pigment epithelium, etc.) or the expressed phenotypes (for example dying cell, cell division, etc.), allowing a totally non invasive imaging in biology. These signals, representing the cellular activity, come from organelles present in the cells, especially mitochondria, and help to an easy interpretation of the condition of the cells.This innovative contrast allows to study retinal degenerative diseases in vitro, such as Age-related Macular Degeneration or Retinitis Pigmentosa. Disease modeling can easily be studied by evaluating the cell response over time, for example after an induced stress on the cell culture (scratch assays). Drug screening for these diseases will also be eased by following the evolution of the dynamic signal produced by the cells after treatment.Finally, the application of Full-Field Optical Coherence Tomography to in vivo retinal imaging permit a non invasive study of the different layers of the retina (from the nerves to the retinal pigment epithelium). It is a technique less complex than those with adaptive optics usually used for in vivo imaging. The stabilisation and signal improvement justify the idea of the future in vivo implementation of dynamic FFOCT for the detection of diseases at early stages, such as Age-related Macular Degeneration or Glaucoma

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

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    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‚ąė√ó5‚ąė field-of-view, without apparent anisoplanatism

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

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    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 √ó 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

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

    No full text
    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

    Optical phase modulation by natural eye movements: application to time-domain FF-OCT image retrieval

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    International audienceEye movements are commonly seen as an obstacle to high-resolution ophthalmic imaging. In this context we study the natural axial movements of the in vivo human eye and show that they can be used to modulate the optical phase and retrieve tomographic images via time-domain full-field optical coherence tomography (TD-FF-OCT). This approach opens a path to a simplified ophthalmic TD-FF-OCT device, operating without the usual piezo motor-camera synchronization. The device demonstrates in vivo human corneal images under the different image retrieval schemes (2-phase and 4-phase) and different exposure times (3.5 ms, 10 ms, 20 ms). Data on eye movements, acquired with a spectral-domain OCT with axial eye tracking (180 B-scans/s), are used to study the influence of ocular motion on the probability of capturing high-signal tomographic images without phase washout. The optimal combinations of camera acquisition speed and amplitude of piezo modulation are proposed and discussed
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