The structural and functional imaging of ophthalmic tissues in cellular level play an important role in the understanding and evaluation of the physiology and pathology of ophthalmic diseases. In this study, we developed a dual-mode full-field optical coherence tomography (FFOCT) that is capable of acquiring label-free cellular images of freshly excised ophthalmic tissues, achieving static contrasts gained from structural refractive index gradients and dynamic contrast induced by endogenous cell motility related to cell functions. Through imaging experiments on both normal and pathological ophthalmic tissues, we show that while the static FFOCT images better reveal the relative stationary cellular structures like nerve fibers and collagens, the dynamic FFOCT images show enhanced contrast of various transparent cells with active intracellular metabolic motions, offering complementary information of major corneal and retinal layers. Our study has shown the dual-mode FFOCT system is a straightforward promising technique for cellular imaging exploration and pathological analysis of ophthalmic tissues.
Eye movements are commonly seen as an obstacle to high-resolution ophthalmic imaging. In this context we study the natural axial movements of 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 different image retrieval schemes (2-phase and 4-phase) and different exposure times (3.5 ms, 10 ms, 20 ms).
KEYWORDS: Optical coherence tomography, In vivo imaging, Real time imaging, Angiography, Eye, Cornea, Confocal microscopy, Microscopes, Imaging systems, Blood circulation
KEYWORDS: In vivo imaging, Retinal scanning, Optical coherence tomography, Wavefronts, 3D image processing, Adaptive optics, Stereoscopy, 3D metrology, Sensors, Eyeglasses
Full-Field Optical Coherence Tomography (FF-OCT) offers aberration independent resolution. This inherent property makes FF-OCT a promising imaging modality for 3D high-resolution retinal imaging. Nevertheless, ocular aberrations affect signal reduction, imposing Adaptive Optics. Here we investigate the best strategy to compensate for ocular aberrations in our FF-OCT setup, in terms of wavefront measurement and correction. The use of wavefront sensorless approach based on the FF-OCT signal level is investigated. Moreover, a strategy of static wavefront correction in a non-conjugated pupil plane and next to the eye’s pupil, just like spectacles, favoring a compact and non-complex AO design, is also investigated. Additionally, the use of wavefront corrector devices such as an adaptive liquid lens (correcting to defocus and astigmatism) and multi-actuator adaptive lenses (correcting up to the 4th order Zernike polynomial) are evaluated. Finally, we expect the implementation of one or a combination of the studied strategies into our FF-OCT setup to lead to the first in-vivo retinal images obtained using AO-assisted FF-OCT, for different retinal layers with enhanced SNR and a 3D high-resolution.
Despite obvious improvements in visualization of the in vivo cornea through the faster imaging speeds and higher axial resolutions, cellular imaging stays unresolvable task for OCT, as en face viewing with a high lateral resolution is required. The latter is possible with FFOCT, a method that relies on a camera, moderate numerical aperture (NA) objectives and an incoherent light source to provide en face images with a micrometer-level resolution. Recently, we for the first time demonstrated the ability of FFOCT to capture images from the in vivo human cornea1. In the current paper we present an extensive study of appearance of healthy in vivo human corneas under FFOCT examination. En face corneal images with a micrometer-level resolution were obtained from the three healthy subjects. For each subject it was possible to acquire images through the entire corneal depth and visualize the epithelium structures, Bowman’s layer, sub-basal nerve plexus (SNP) fibers, anterior, middle and posterior stroma, endothelial cells with nuclei. Dimensions and densities of the structures visible with FFOCT, are in agreement with those seen by other cornea imaging methods. Cellular-level details in the images obtained together with the relatively large field-of-view (FOV) and contactless way of imaging make this device a promising candidate for becoming a new tool in ophthalmological diagnostics.
We show for the first time FF-OCT combined with SD-OCT for real time matching of the optical path length of FF-OCT to demonstrate high resolution en face retinal imaging.
Optical imaging usually suffers from aberrations that are induced by various structures when imaging biological samples. Usually aberrations degrade the imaging system performances by broadening the point spread function (PSF). Unexpectedly we show that in spatially incoherent interferometry like full-filed optical coherence tomography (FFOCT), the system PSF width is almost insensitive to aberrations. Instead of considering the PSF of a classical imaging system such as a microscope, we specifically pay attention to the system PSF of interferometric imaging systems for which an undistorted wavefront from a reference beam interferes with the distorted wavefront of the object beam. By comparing the cases of scanning OCT with spatially coherent illumination, wide-field OCT with spatially coherent illumination and FFOCT with spatially incoherent illumination, we found that in FFOCT with spatially incoherent illumination the system PSF width is almost independent of the aberrations and only its amplitude varies. This is demonstrated by theoretical analysis as well as numerical calculations for different aberrations, and confirmed by experiments with a FFOCT system. It is the first time to the best of our knowledge that such specific merit of incoherent illumination in FFOCT has been demonstrated. Based on this, the signal level is used as metric in our adaptive optics FFOCT system for retinal imaging. Only the main aberrations (defocus and astigmatism) that are dominating in eye are corrected to improve the signal to noise ratio and the high order aberrations are skipped. This would increase the correction speed thus reducing the imaging time.
Adaptive optics full-filed OCT (FFOCT) with a transmissive liquid crystal spatial light modulator
(LCSLM) as wavefront corrector is used without strict plane conjugation for low order aberrations
corrections. We validated experimentally that FFOCT resolution is independent of aberrations and
only reduce the signal level. A signal based sensorless algorithm was thus applied for wavefront
distortion compensation. Image quality improvements by the wavefront sensorless control of the
LCSLM were evaluated on in vitro samples. By replacing the FFOCT sample arm objective with an
artificial eye used to train ophthalmologists, adaptive optics retinal imaging was achieved. In vivo
experiments using a liquid lens to correct focus and astigmatism are underway.
We describe a simple and compact full-field optical coherence tomography (FFOCT) setup coupled to a transmissive liquid crystal spatial light modulator (LCSLM) to induce or correct aberrations. To reduce the system complexity, strict pupil conjugation was abandoned because low-order aberrations are often dominant. We experimentally confirmed a recent theoretical and experimental demonstration that the image resolution was almost insensitive to aberrations that mostly induce a reduction of the signal level. As a consequence, an image-based algorithm was applied for the optimization process by using the FFOCT image intensity as the metric. Aberration corrections were demonstrated with both an USAF resolution target and biological samples for LCSLM-induced and sample-induced wavefront distortions.
An aberrated imaging system PSF is broadened; this broadening is responsible of the blurring of the images. A lot of effort has been carried out to correct the effects of aberrations on OCT images for eye examination or biological samples. We have worked on quantifying the effect of geometrical aberrations on Full-Field OCT images and found that there is mostly no loss of resolution but a decrease of the signal level. This is obviously why we use these signals as metric to correct the wavefront distortion. Moreover we found that this absence of blurring, which is due to the fact that we record the dot product of a diffraction limited reference signal and the distorted sample signal, is specific to the use of an incoherent illumination and did not show up with OCT approaches that use spatially coherent sources. More precisely the loss in signal is roughly proportional to the square root of the Strehl ratio: for example, a Strehl ratio of 1/9, which is considered to give a low quality image, would only be 1/3 in Full-Field OCT while keeping the sharpness of the image. Using both an USAF resolution target and a transmissive SLM we have demonstrated this unique feature of sharpness conservation. It was also confirmed by using biological samples. We think that we can thus restrict the aberration corrections in eye examination to the main aberrations (e.g. focus and astigmatism) that will increase the speed of the correction.
A Full-Field OCT (FFOCT) setup coupled to a compact transmissive liquid crystal spatial light modulator (LCSLM) is used to induce or correct aberrations and simulate eye examinations. To reduce the system complexity, strict pupil conjugation was abandoned. During our work on quantifying the effect of geometrical aberrations on FFOCT images, we found that the image resolution is almost insensitive to aberrations. Indeed if the object channel PSF is distorted, its interference with the reference channel conserves the main feature of an unperturbed PSF with only a reduction of the signal level. This unique behavior is specific to the use of a spatially incoherent illumination. Based on this, the FFOCT image intensity was used as the metric for our wavefront sensorless correction. Aberration correction was first conducted on an USAF resolution target with the LSCLM as both aberration generator and corrector. A random aberration mask was induced, and the low-order Zernike Modes were corrected sequentially according to the intensity metric function optimization. A Ficus leaf and a fixed mouse brain tissue slice were also imaged to demonstrate the correction of sample self-induced wavefront distortions. After optimization, more structured information appears for the leaf imaging. And the high-signal fiber-like myelin fiber structures were resolved much more clearly after the whole correction process for mouse brain imaging. Our experiment shows the potential of this compact AO-FFOCT system for aberration correction imaging. This preliminary approach that simulates eyes aberrations correction also opens the path to a simple implementation of FFOCT adaptive optics for retinal examinations.
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