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Scott M McClintic,1 Dae Yu Kim,2 Jeff Fingler,3 Susan Garcia,4 Robert J Zawadzki,4 Lawrence S Morse,4 Susanna S Park,4 Scott E Fraser,3 John S Werner,4 Jason P Ruggiero,5 Daniel M Schwartz6 1Department of Ophthalmology, Casey Eye Institute, Oregon Health and Science University, Portland, OR, USA; 2Beckman Laser Institute Korea and Department of Biomedical Engineering, Dankook University, Cheonan, South Korea; 3Translational Imaging Center, University of Southern California, Los Angeles, CA, 4Department of Ophthalmology and Vision Science, University of California Davis, Sacramento, CA, 5Retina Associates, PC, Winchester, VA, 6Department of Ophthalmology and Vision Science, University of California San Francisco, San Francisco, CA, USA Purpose: To demonstrate the use of phase-variance optical coherence tomography (PV-OCT) angiography for detection of pigment epithelial detachment (PED) vascularization in age-related macular degeneration (AMD). Patients and methods: Patients with PEDs and exudative AMD were evaluated by the Retina Services at the University of California, Davis, and the University of California, San Francisco. Each subject underwent fluorescein angiography and structural optical coherence tomography (OCT). Phase-variance OCT analysis was used to create angiographic images of the retinal and choroidal vasculature. PV-OCT-generated B-scans were superimposed on structural OCT B-scans to allow easy identification of perfused vascular structures. Results: Three patients with vascularized PEDs were imaged with PV-OCT, and each was found to have a vascular signal extending from the choroid into the hyperreflective substance of the PED. Two patients who had no evidence of PED vascularization on fluorescein angiography did not have vascular signals within their PEDs on PV-OCT. Conclusion: Structural OCT and PV-OCT images can be combined to create composite B-scans that offer high-resolution views of the retinal tissue along with dynamic vascular visualization. This technique offers a fast, noninvasive method for detecting vascularization of PEDs in AMD and may aid in the early detection of neovascular disease. Keywords: OCT, imaging, retina

We describe the design and performance of a recently implemented retinal imaging system for the human eye that combines adaptive optics (AO) with spectral domain optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO). The AO-OCT-SLO system simultaneously acquires SLO frames and OCT B-scans at 60 Hz with an OCT volume acquisition scan rate of 0.24 Hz. The SLO images are used to correct for eye motion during the registration of OCT B-scans. Key optical design considerations are discussed including: minimizing system aberrations through the use of off-axis relay telescopes; choice of telescope magnification based on pupil plane requirements and restrictions; and the use of dichroic beam splitters to separate and re-combine OCT and SLO beams around the nonshared horizontal scanning mirrors. We include an analysis of closed-loop AO correction on a model eye and compare these findings with system performance in vivo. The 2D and 3D OCT scans included in this work demonstrate the ability of this system to laterally and axially resolve individual cone photoreceptors, while the corresponding SLO images show the en face mosaics at the photoreceptor layer showing rods and cones. Images from both healthy and diseased retina are presented.

We tested and compared the capability of multiple optical coherence tomography (OCT) angiography methods: phase variance, amplitude decorrelation and speckle variance, with application of the split spectrum technique, to image the choroiretinal complex of the human eye. To test the possibility of OCT imaging stability improvement we utilized a real-time tracking scanning laser ophthalmoscopy (TSLO) system combined with a swept source OCT setup. In addition, we implemented a post- processing volume averaging method for improved angiographic image quality and reduction of motion artifacts. The OCT system operated at the central wavelength of 1040nm to enable sufficient depth penetration into the choroid. Imaging was performed in the eyes of healthy volunteers and patients diagnosed with age-related macular degeneration.

We demonstrate the feasibility of our newly developed phase stabilized high-speed (100 kHz A-scans/s) 1 μm sweptsource optical coherence tomography (SSOCT) system with the phase-variance based motion contrast method for visualization of human chorioretinal complex microcirculation. Compared to our previously reported spectral domain (spectrometer based) phase-variance (pv)-SDOCT system it has advantages of higher sensitivity, reduced fringe washout for high blood flow speeds and deeper penetration in choroid. High phase stability SSOCT imaging was achieved by using a computationally efficient phase stabilization approach. This process does not require additional calibration hardware and complex numerical procedures. Our phase stabilization method is simple and can be employed in a variety of SS-OCT systems. Examples of vasculature in the chorioretinal complex imaged by pv-SSOCT is presented and compared to retinal images of the same volunteers acquired with fluorescein angiography (FA) and indocyanine green angiography (ICGA).

We present a new design for a reflective afocal AO-OCT retinal imaging system. The optical performance of this instrument is compared to our previous multimodal AO-OCT/AO-SLO retinal imaging system. The feasibility of new instrumentation for improved visualization of microscopic retinal structures will be discussed. Examples of images acquired with this new AO-OCT instrument will be presented. © 2013 Copyright SPIE.

We present in vivo noninvasive retinal and choroidal perfusion maps with phase-variance optical coherence tomography (pvOCT). We acquired a pvOCT volumetric data set of a normal subject and visualized blood circulation in the retina and the choroid. En face projection views of the retina as well as the choroid were generated from a manually segmented volumetric data set. In addition, the processed pvOCT images were compared to current standard imaging modalities used for retinal and choroidal vasculature visualization in clinical settings, including fluorescein angiography (FA) and indocyanine green angiography (ICGA). © 2013 Copyright SPIE.

We present a high-speed complex conjugate resolved 1 μm swept source optical coherence tomography [SS-OCT] system using coherence revival of the light source for clinical imaging of the anterior segment of the eye. High-speed of 100,000 A-scans/sec and 1 μm imaging window of OCT permits dense 3D imaging of the anterior segment, minimizing the influence of motion artifacts and deep penetration of images for topographic analysis. The swept laser performance with internal clocking was adapted to achieve extended imaging depth requirements. The feasibility of our instrument for visualization of the anterior segment of patients with the Boston Keratoprosthesis (KPro) was discussed. The relations between of the KPro and the surrounding tissue were also demonstrated. © 2013 Copyright SPIE.

Recent progress in retinal image acquisition techniques, including optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO), combined with improved performance of adaptive optics (AO) instrumentation, has resulted in improvement in the quality of in vivo images of cellular structures in the outer layers of the human retina. Despite the significant progress in imaging cone and rod photoreceptor mosaics, visualization of cellular structures in the inner retina has been achieved only with extrinsic contrast agents that have not been approved for use with humans. In this paper we describe the main limiting factors in visualizing inner retinal cells and the methods we implemented to reduce their effects on images acquired with AO-OCT. These include improving the system point spread function (AO performance), monitoring of motion artifacts (retinal motion tracking), and speckle pattern reduction (temporal and spatial averaging). Results of imaging inner retinal morphology and the improvement offered by the new UC Davis AOOCT system with spatio-temporal image averaging are presented. © 2012 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).

We evaluate methods to visualize human retinal micro-circulation in vivo by standard intensity-based optical coherence tomography (OCT), speckle-variance optical coherence tomography (svOCT), and phase-variance optical coherence tomography (pvOCT). En face projection views created from the same volumetric data set of the human retina using all three data processing methods are created and compared. Additionally we used support vector machine (SVM) based semi-automatic segmentation to generate en face projection views of individual retinal layers. The layers include: first, the whole inner retina (from the nerve fiber layer to the outer nuclear layer), and second, from the ganglion cell layer to the outer nuclear layer. Finally, we compare the retinal vasculature images processed from the three OCT techniques and fluorescein angiography (FA). © 2012 SPIE.

Current developments in cellular resolution in-vivo retinal imaging systems at the UC Davis will be presented. Instrumentation developments include the combination of adaptive optics with optical coherence tomography and scanning laser ophthalmoscopy. © 2011 Optical Society of America.

We present an evaluation of two different configurations of the ALPAO 97-actuator membrane magnetic deformable mirror for wavefront correction, using either 7 or 9 actuators across the eye pupil. These tests included monitoring of aberration correction for a human subject. This AO-sub system is part of the UC Davis high-resolution human retinal imaging adaptive optics - optical coherence tomography (AO-OCT) instrument. AO-OCT allows the three-dimensional (3D) visualization of different retinal structures in vivo with high volumetric resolution (3×3×3μm 3). © 2011 Photonics Society of Poland.

Nowadays in ophthalmologic practice several commercial instruments are available to image patient retinas in vivo. Many modern fundus cameras and confocal scanning laser ophthalmoscopes allow acquisition of two dimensional en face images of the retina with both back reflected as well as fluorescent light. Additionally, optical coherence tomography systems allow non-invasive probing of three-dimensional retinal morphology. For all of these instruments the available lateral resolution is limited by optical quality of the human eye used as the imaging objective. To improve lateral resolution and achieve diffraction-limited imaging, adaptive optics (AO) can be implemented with any of these imaging systems to correct both static and dynamic aberrations inherent in human eyes. Most of the wavefront correctors used previously in AO systems have limited dynamic range and an insufficient number of actuators to achieve diffraction-limited correction of most human eyes. Thus, additional corrections were necessary, either by trial lenses or additional deformable mirrors (DMs). The UC Davis AO flood-illuminated fundus camera system described in this paper has been previously used to acquire in vivo images of the photoreceptor mosaic and for psychophysical studies on normal and diseased retinas. These results were acquired using a DM manufactured by Litton ITEK (DM109), which has 109 actuators arranged in a hexagonal array below a continuous front-surface mirror. It has an approximate surface actuator stroke of ±2μm. Here we present results with a new hi-speed magnetic DM manufactured by ALPAO (DM97, voice coil technology), which has 97 actuators and similar inter-actuator stroke (>3μm, mirror surface) but much higher low-order aberration correction (defocus stroke of at least ±30μm) than the previous one. In this paper we report results of testing performance of the ALPAO DM for the correction of human eye aberrations. Additionally changes made to our AO flood illuminated system are presented along with images of the model eye retina and in-vivo human retina acquired with this system. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).

PurposeThe purpose of this study is to evaluate the macular morphological changes associated with idiopathic epiretinal membrane (iERM) using high-resolution Fourier-domain optical coherence tomography (FD-OCT), as they correlate with visual acuity and microperimetry (MP-1).MethodsIn all, 24 eyes (19 subjects) with iERM were imaged prospectively using FD-OCT with axial resolution of 4.5 μm and transverse resolution of 10 to 15 μm. MP-1 and Stratus OCT were carried out in a subset of eyes.ResultsThe mean log of the minimum angle of resolution best-corrected visual acuity (BCVA) was 0.18±0.16 (range: -0.08 to 0.48, Snellen equivalent 20/15(-1) to 20/60). ERM was visualized in all 24 eyes with FD-OCT and in 17 eyes (85%) of 20 eyes imaged with Stratus OCT. Although BCVA correlated with macular thickening in the central 1 mm sub-field of the Stratus ETDRS (P=0.0005) and macular volume (central 3 mm area) on FD-OCT (P

We present high-speed Fourier-domain optical coherence tomography (Fd-OCT) with the phase variance based motion contrast method for visualizing retinal micro-circulation in vivo. This technique allows non-invasive visualization of a two-dimensional retinal perfusion map and concurrent volumetric morphology of retinal microvasculature with high sensitivity. The high-speed acquisition rate at 125kHz A-scans enables reduction of motion artifacts with increased scanning area if compared to previously reported results. Several scanning schemes with different sampling densities and scanning areas are evaluated to find optimal parameters for in vivo imaging. In order to evaluate this technique, we compare OCT micro-capillary imaging using the phase variance technique with fundus fluorescein angiography (FA). Additionally, volumetric visualization of blood flow for a normal subject is presented. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).

Eye movements present during acquisition of a retinal image with optical coherence tomography (OCT) introduce motion artifacts into the image, complicating analysis and registration. This effect is especially pronounced in highresolution data sets acquired with adaptive optics (AO)-OCT instruments. Several retinal tracking systems have been introduced to correct retinal motion during data acquisition. We present a method for correcting motion artifacts in AOOCT volume data after acquisition using simultaneously captured adaptive optics-scanning laser ophthalmoscope (AOSLO) images. We extract transverse eye motion data from the AO-SLO images, assign a motion adjustment vector to each AO-OCT A-scan, and re-sample from the scattered data back onto a regular grid. The corrected volume data improve the accuracy of quantitative analyses of microscopic structures. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).

Inner and outer retinal morphology were quantified in vivo for 6 nonglaucomatous and 10 glaucomatous optic neuropathy patients. Custom, ultrahigh-resolution imaging modalities were used to evaluate segmented retinal layer thickness in 3D volumes (Fourier-domain optical coherence tomography), cone photoreceptor density (adaptive optics fundus camera), and the length of inner and outer segments of cone photoreceptors (adaptive optics-optical coherence tomography). Quantitative comparisons were made with age-matched controls, or by comparing affected and nonaffected retinal areas defined by changes in visual fields. The integrity of outer retinal layers on optical coherence tomography B-scans and density of cone photoreceptors were correlated with visual field sensitivity at corresponding retinal locations following reductions in inner retinal thickness. The photoreceptor outer segments were shorter and exhibited greater variability in retinal areas associated with visual field losses compared with normal or less affected areas of the same patient's visual field. These results demonstrate that nonglaucomatous and glaucomatous optic neuropathies are associated with outer retinal changes following long-term inner retinal pathology.

An accurate solid eye model (with volumetric retinal morphology) has many applications in the field of ophthalmology, including evaluation of ophthalmic instruments and optometry/ophthalmology training. We present a method that uses volumetric OCT retinal data sets to produce an anatomically correct representation of three-dimensional (3D) retinal layers. This information is exported to a laser scan system to re-create it within solid eye retinal morphology of the eye used in OCT testing. The solid optical model eye is constructed from PMMA acrylic, with equivalent optical power to that of the human eye (∼58D). Additionally we tested a water bath eye model from Eyetech Ltd. with a customized retina consisting of five layers of ∼60 μm thick biaxial polypropylene film and hot melt rubber adhesive. © 2010 SPIE.

We describe results of retinal imaging with a novel instrument that combines adaptive optics - Fourier-domain optical coherence tomography (AO-OCT) with an adaptive optics scanning laser ophthalmoscope (AO-SLO). One of the benefits of combining Fd-OCT with SLO includes automatic co-registration between the two imaging modalities and the potential for correcting lateral and transversal eye motion resulting in motion artifact-free volumetric retinal imaging. Additionally this allows for direct comparison between retinal structures that can be imaged with both modalities (e.g., photoreceptor mosaics or microvasculature maps). This dual imaging modality could provide insight into some retinal properties that could not be accessed by a single imaging system. Additionally, extension of OCT and SLO beyond structural imaging may open new avenues for diagnostics and testing in ophthalmology. In particular, non-invasive vasculature mapping with these modalities holds promise of replacing fluorescein angiography in vascular identification. Several new improvements of our system are described, including results of testing a novel 97-actuator deformable mirror and AO-SLO light intensity modulation. © 2010 SPIE.

We evaluated several, previously published, complex conjugate artifact removal methods and algorithms that have been proposed for Fourier domain optical coherence tomography (Fd-OCT). To ensure comparable conditions, only one OCT system was used, but with modified data acquisition schemes, depending on the requirements of each method/algorithm. This limited our evaluation to single spectrometer based Fd-OCT approaches. The suppression ratio of complex conjugate artifact images using a paperboard is assessed for all tested methods. Several other metrics are also used for comparison, including a list of additional hardware requirements (beyond standard Fd-OCT components) and data acquisition schemes. Finally, in vivo human finger pad and nail images are presented for comparison to the standard Fd- OCT images and full-range images. © 2010 Copyright SPIE - The International Society for Optical Engineering.

Imaging the structure and correlating it with the biochemical content of the retina holds promise for fundamental research and for clinical applications. Optical coherence tomography (OCT) is commonly used to image the 3D structure of the retina and while the added functionality of biochemical analysis afforded by Raman scattering could provide critical molecular signatures for clinicians and researchers, there are many technical challenges to combining these imaging modalities. We present an ex vivo OCT microscope combined with Raman spectroscopy capable of collecting morphological and molecular information about a sample simultaneously. The combined instrument will be used to investigate remaining technical challenges to combine these imaging modalities, such as the laser power levels needed to achieve a Raman signal above the noise level without damaging the sample. © 2009.

We describe a novel instrument that combines adaptive optics - Fourier-domain optical coherence tomography (AO-OCT) with an adaptive optics scanning laser ophthalmoscope (AO-SLO). Both systems share a common AO sub-system and vertical scanner to permit simultaneous acquisition of retinal images from both OCT and SLO. One of the benefits of combining OCT with SLO includes automatic co-registration between the two imaging modalities and potential for correcting lateral and transversal eye motion resulting in motion artifact-free volumetric retinal imaging. Results of using this system for eye model imaging are presented. Feasibility for clinical application is briefly discussed as well as potential further improvements of the current system. © 2009 SPIE.

ObjectivesTo investigate retinal microstructure of patients affected with malattia leventinese (MLVT) and mutation in the EFEMP1 gene using high-resolution optical coherence tomography (OCT).MethodsPatients diagnosed with MLVT received a comprehensive eye exam, full-field and multifocal electroretinogram testing and imaging with a high-resolution Fourier domain OCT (Fd-OCT, UC Davis Medical Center, Davis, USA; axial resolution: 4.5 microm, acquisition speed: 9 frames s(-1), 1000 A scans s(-1)) combined with a flexible scanning head (Bioptigen Inc. Durham, NC, USA).ResultsTwo related patients aged 30 and 60 years, with MLVT and identified c.R345W mutation in the EFEMP1 gene, were tested. Mother and daughter showed a variable phenotype with reduced vision function in the younger patient, whereas the mother had a 'form frustre'. Fd-OCT revealed extensive or focal sub-retinal pigment epithelium (RPE) deposits, separation of RPE and Bruch's membrane, and disruption of the photoreceptor outer and inner segment layers. No outer retinal changes were visible outside areas with sub-RPE deposits.ConclusionRetinal structure in EFEMP1 retinal dystrophy is reflected by morphological changes within the RPE/Bruch's membrane complex with accumulation of sub-RPE material associated with disrupted photoreceptor integrity. The pattern of microstructural retinal abnormalities is similar but with a different extent in patients with variable phenotypes.

Recent developments in adaptive optics - optical coherence tomography (AO-OCT) allow for ultra-high isotropic resolution imaging of the in-vivo retina, offering unprecedented insight into its volumetric microscopic and cellular structures. In addition to this promising achievement, the clinical impact and application of this technology still needs to be explored. This includes assessment of limitations and challenges for existing as well as future AO-OCT systems, especially in the context of potential transfer of this technology from an optical bench to a portable imaging system. To address these issues we will describe our current AO-UHR-OCT focusing on its sub-components, as well as application for clinical imaging. Additionally, we describe some directions for future development of our AO-OCT instrument that would improve its clinical utility including: new compact AO-OCT design, new improved AO sub-system (extreme AO), and new generations of Fourier-domain-OCT. © 2008 SPIE.

Ultra-high-resolution (UHR) optical coherence tomography (OCT) imaging technique is using adaptive optics, to perform better better treatment and monitoring of retinal diseases. Researchers have developed a new generation of adaptive optics (AO) UHR OCT (AO-UHR-OCT) instrument that combines an ultrabroadband spectral light source and an achromatizing element, to increase measured axial resolution to around 3.5 μm. The measured axial resolution is increased without reducing lateral resolution and image quality. The new instrument uses an AO system in the spectral/Fourier domain OCT sample arm, to facilitate correction of a patient's ocular aberrations. The detection channel of the OCT spectrometer acquires spectral fringes of light, back-scattered from the retina, along with light from the reference arm.

Ultra-high isotropic resolution imaging of retinal structures was made possible with an adaptive optics system using dual deformable mirrors and a Fourier-domain optical coherence tomography (Fd-OCT) system with correction for longitudinal chromatic aberration. This system was used to image microscopic retinal structures of healthy as well as diseased retinas in vivo. The improved resolution and contrast enhanced visualization of morphological structures in the retina can be clearly seen. The benefits of this instrument are apparent from comparison of new images with those acquired using a previous generation AO-OCT instrument. Big change in the appearance of speckle field (reduction in speckle size) can be observed as well. Additionally, further improvements in volumetric data acquisition and image representation will be discussed. This includes creation of large Field of View (FOV) AO-OCT volume from multiple sub-volumes and its visualization. Also techniques and results of reducing speckle contrast by averaging multiple B-scans will be presented.

Adaptive optics (AO) and optical coherence tomography (OCT) are powerful imaging modalities that, when combined, can provide high-resolution, 3-D images of the retina. The AO-OCT system at UC Davis has been under development for 2 years and has demonstrated the utility of this technology for microscopic, volumetric, in vivo retinal imaging. The current system uses a bimorph deformable mirror (DM) made by AOptix Technologies, Inc. for low-order, high-stroke correction and a 140-actuator mirco-electrical-mechanical-system (MEMS) DM made by Boston Micromachines Corporation for high-order correction. We present our on-going characterization of AO system performance. The AO-OCT system typically has residual wavefront error of 100 nm rms. The correctable error in the system is dominated by low-order error that we believe is introduced by aliasing in the control loop. Careful characterization of the AO system will lead to improved performance and inform the design of future systems.

We describe a novel instrument capable of acquiring, simultaneously, adaptive optics enhanced scanning laser ophthalmoscopy and optical coherence tomography (OCT) images of the human cone mosaic in vivo. The OCT system is based on transversal scanning of the sample with a line scan rate of 14 kHz, approximately 20 times faster than a previously reported instrument. We demonstrate the capability of this instrument with the measurement of the human cone spacing in perifoveal retina.

The acquisition speed of current FD-OCT (Fourier Domain - Optical Coherence Tomography) instruments allows rapid screening of three-dimensional (3D) volumes of human retinas in clinical settings. To take advantage of this ability requires software used by physicians to be capable of displaying and accessing volumetric data as well as supporting post processing in order to access important quantitative information such as thickness maps and segmented volumes. We describe our clinical FD-OCT system used to acquire 3D data from the human retina over the macula and optic nerve head. B-scans are registered to remove motion artifacts and post-processed with customized 3D visualization and analysis software. Our analysis software includes standard 3D visualization techniques along with a machine learning support vector machine (SVM) algorithm that allows a user to semi-automatically segment different retinal structures and layers. Our program makes possible measurements of the retinal layer thickness as well as volumes of structures of interest, despite the presence of noise and structural deformations associated with retinal pathology. Our software has been tested successfully in clinical settings for its efficacy in assessing 3D retinal structures in healthy as well as diseased cases. Our tool facilitates diagnosis and treatment monitoring of retinal diseases.

We report first observations of the three-dimensional morphology of cone photoreceptors in the living human retina. Images were acquired with a high-speed adaptive optics (AO) spectral domain optical coherence tomography (SD-OCT) camera. The AO system consists of a Shack-Hartmann wavefront sensor and bimorph deformable mirror (AOptix) that measure and correct the ocular and system aberrations at a closed-loop rate of 12 Hz. Unlike previous AO-OCT and AO-SLO instruments, the bimorph mirror was strategically positioned between the XY mechanical scanners and the subject's eye so as to avoid beam distortion at the pupil plane, which is created when the mirror compensates for the refractive error of the eye. This new configuration is evaluated empirically and with commercial ray tracing software. The SD-OCT system consists of a superluminescent diode and a 512 pixel line scan charge-coupled device (CCD) that acquires 75,000 A-scans/sec. This rate is more than two times faster than that previously reported. Retina motion artifiacts were minimized by quickly acquiring small volume images of the retina with and without AO compensation. Camera sensitivity was sufficient to detect reflections from all major retinal layers. The distribution of bright spots observed within C-scans at the inner segment / outer segment (IS/OS) junction and at the posterior tips of the OS were found to be highly correlated with one another and with the expected cone spacing. No correlation was found between the IS/OS junction and either the plexiform layers or the layers immediately behind the OS posterior tips.

Adaptive optics - optical coherence tomography (AO-OCT) has the potential to improve lateral resolution for OCT retinal imaging. Several reports have already described the successful combination of AO with a scanning confocal Fourier-domain OCT instrument to permit real-time three-dimensional (3D) imaging with high resolution (in all three dimensions). One of the key components that sets the performance limit of AO is the wavefront corrector. Several different wavefront correctors have been used in AO-OCT systems so far. In this paper we compare two commercially available wavefront correctors: an AOptix Bimorph deformable mirror (DM) and a Boston Micromachines Micro-Electro Mechanical System (MEMS) DM (used for the first time in an AO-OCT system). To simplify the analysis, we tested their performance for the correction of low-amplitude high-order aberrations (with minimal defocus and astigmatism). Results were obtained with an AO-OCT instrument constructed at UC Davis that combines state-of-the-art Fourier-domain OCT and an AO design to allow simultaneous testing of both mirrors without the need to modify the optical system.

Adaptive optics (AO) is used for improving lateral resolution in opthalmic Optical-Coherence-Tomography (OCT) to correct for the aberrations in the human eye. AO works by measuring aberrations using a wavefront sensor and applying this information to the control of a deformable mirror in a closed loop, thereby compensating for the measured aberrations. This instrument combines the advantages of FD-OCT and advanced microtechnology for AO, using 35-element bimorph deformable mirror. AO system can effectively compensate low-order aberrations and maintain low rms wavefront error during closed-loop operations.

We have combined Fourier-domain optical coherence tomography (OCT) with a closed-loop Adaptive Optics (AO) system. The AO-OCT instrument has been used for in vivo retinal imaging. High-lateral resolution of our AO-OCT system allows visualization of the microscopic retinal structures not accessible by standard OCT instruments. © 2005 SPIE and OSA.

We built a Fourier domain optical coherence tomography (FD-OCT) system using a line scan CCD camera that allows real time data display and acquisition. This instrument is able to produce 2D B-scans as well as 3D data sets with human subjects in vivo in clinical settings. In this paper we analyze the influence of varying exposure times of the CCD detector on image quality. Sensitivity values derived from theoretical predictions have been compared with measurements (obtained with mirrors and neutral density filters placed in both interferometer arms). The results of these experiments, discussion about differences between sensitivity values, potential sources of discrepancies, and recommendations for optimal exposure times will be described in this paper. A short discussion of observed artifacts as well as possible ways to remove them is presented. The influence of relative retinal position with respect to reference mirror position will also be described.

Retinal dystrophy in Bardet-Biedl Syndrome (BBS) is caused by defective genes that are expressed within ciliated cells such as photoreceptors. The purpose of this study was to characterize and compare the retinal structure and lamination of two groups of patients, carrying mutations in BBS1 or BBS10. Eight patients with BBS (ages 11.9-28.5 years) and mutations in BBS1 (4/8) or BBS10 (4/8) were tested. A high-resolution hand-held probe Fourier-domain optical coherence tomography system (Fd-OCT) was used for retinal image acquisition. Macular scans were evaluated with respect to structure, retinal layering and photoreceptor integrity. Micro-structural in-vivo analysis showed abnormalities within retinal layers but preserved retinal lamination. Photoreceptor integrity was disrupted in all patients. Macular scans from patients with BBS10 mutations most often showed 'deposits' adjacent and anterior to Bruch's membrane. Age, genotype and presence of macular changes did not correlate with the structural changes observed. Retinal dystrophy in BBS is reflected by major changes in the outer retinal layers. This is the first report of in-vivo micro-structural analysis of retinal layers in patients with BBS. Mutations in different BBS genes seem to be associated with similar micro-structural changes in retinal layers. [ABSTRACT FROM AUTHOR]

A full-field swept-source optical coherence tomography (FF-SS-OCT) for in vivo murine retinal imaging is demonstrated. The on-axis FF-SS-OCT system was built in a Mach-Zehnder interferometer configuration employing a tunable laser source with an adjustable sweep rate and sweep range in conjunction with a fast 2D-CMOS camera. A large field retinal (coherent) illumination was accomplished using an imaging interface comprised of a short-focal length imaging lens and a contact lens. The magnification between the camera and retina (spatial sampling) was appropriately chosen to record the microscopic structural features of the retina in the image. A pupil stop was employed in the detection path to reject unwanted backscattering from the mouse eye and other sources and limit aberrations distorting the retinal images. In vivo mouse retinal imaging was performed at a sweep rate of 150 Hz to acquire volumes unaffected by the system vibrations, which predominated at lower frequencies. Operating the FF-SS-OCT at this speed yielded an effective axial scan rate of 20 million A-scans/s and a field of view of 820 × 410 µm (24.12° × 12.06°). High-quality retinal B-scans and enface images of the retina were obtained with the SS-FF-OCT, revealing all major retinal layers and vascular plexuses.

The results of depth-resolved multi-contrast in vivo mouse choroidal imaging using a polarization-diversity optical coherence tomography (PD-OCT) system are presented. A selectively chosen depth of focus that was fine-tuned with a sensorless adaptive optics technique and a simple segmentation based on the degree of polarization uniformity signal visualizes the detailed features of a mouse choroid from the OCT angiography images. A comprehensive image analysis of the choroid revealed the distinctive pathological characteristics of the laser-induced choroidal neovascularization mouse.

High-speed, phase contrast retinal and blood flow imaging using an adaptive optics partially confocal multi-line ophthalmosocope (AO-pcMLO) is described. It allows for simultaneous confocal and phase contrast imaging with various directional multi-line illumination by using a single 2D camera and a digital micromirror device (DMD). Both vertical and horizontal line illumination directions were tested, for photoreceptor and vascular imaging. The phase contrast imaging provided improved visualization of retinal structures such as cone inner segments, vessel walls and red blood cells with images being acquired at frame rates up to 500 Hz. Blood flow velocities of small vessels (<40 µ m in diameter) were measured using kymographs for capillaries and cross-correlation between subsequent images for arterioles or venules. Cardiac-related pulsatile patterns were observed with normal resting heart-beat rate, and instantaneous blood flow velocities from 0.7 to 20 mm/s were measured., Competing Interests: The authors declare no conflicts of interest., (© 2024 Optica Publishing Group.)

Glaucoma is a group of eye diseases characterized by the thinning of the retinal nerve fiber layer (RNFL), which is primarily caused by the progressive death of retinal ganglion cells (RGCs). Precise monitoring of these changes at a cellular resolution in living eyes is significant for glaucoma research. In this study, we aimed to assess the effectiveness of temporal speckle averaging optical coherence tomography (TSA-OCT) and dynamic OCT (dOCT) in examining the static and potential dynamic properties of RGCs and RNFL in living mouse eyes. We evaluated parameters such as RNFL thickness and possible dynamics, as well as compared the ganglion cell layer (GCL) soma density obtained from in vivo OCT, fluorescence scanning laser ophthalmoscopy (SLO), and ex vivo histology., Competing Interests: The authors declare no conflicts of interest., (© 2023 Optica Publishing Group.)

A high-speed, adaptive optics partially confocal multi-spot ophthalmoscope (AO-pcMSO) using a digital micromirror device (DMD) in the illumination channel and a fast 2D CMOS camera is described. The camera is synchronized with the DMD allowing projection of multiple, simultaneous AO-corrected spots onto the human retina. Spatial filtering on each raw retinal image before reconstruction works as an array virtual pinholes. A frame acquisition rate of 250 fps is achieved by applying this parallel projection scheme. The contrast improves by 2-3 fold when compared to a standard flood illumination architecture. Partially confocal images of the human retina show cone and rod photoreceptors over a range of retinal eccentricities.

Optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO) are imaging technologies invented in the 1980s that have revolutionized the field of in vivo retinal diagnostics and are now commonly used in ophthalmology clinics as well as in vision science research. Adaptive optics (AO) technology enables high-fidelity correction of ocular aberrations, resulting in improved resolution and sensitivity for both SLO and OCT systems. The potential of gathering multi-modal cellular-resolution information in a single instrument is of great interest to the ophthalmic imaging community. Although similar instruments have been developed for imaging the human retina, developing such a system for mice will benefit basic science research and should help with further dissemination of AO technology. Here, we present our work integrating OCT into an existing mouse retinal AO-SLO system, resulting in a multi-modal AO-enhanced imaging system of the living mouse eye. The new system allows either independent or simultaneous data acquisition of AO-SLO and AO-OCT, depending on the requirements of specific scientific experiments. The system allows a data acquisition speed of 200 kHz A-scans/pixel rate for OCT and SLO, respectively. It offers ∼6 µm axial resolution for AO-OCT and a ∼1 µm lateral resolution for AO-SLO-OCT imaging., Competing Interests: The authors declare that there are no conflicts of interest., (© 2022 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

Purpose: To perform in vivo evaluation of the structural morphology and vascular plexuses of the neurosensory retina and choroid across vertebrate species using swept-source optical coherence tomography (SS-OCT) and SS-OCT angiography (SS-OCTA) imaging., Methods: A custom-built SS-OCT system with an incorporated flexible imaging arm was used to acquire the three-dimensional (3D) retinal OCT and vascular OCTA data of five different vertebrates: a mouse (C57BL/6J), a rat (Long Evans), a gray short-tailed opossum (Monodelphis domestica), a white sturgeon (Acipenser transmontanus), and a great horned owl (Bubo virginianus)., Results: In vivo structural morphology of the retina and choroid, as well as en face OCTA images of retinal and choroidal vasculature of all species were generated. The retinal morphology and vascular plexuses were similar between rat and mouse, whereas distinct choroidal and paired superficial vessels were observed in the opossum retina. The retinal and vascular structure of the sturgeon, as well as the pecten oculi and overlying the avascular and choroidal vasculature in the owl retina are reported in vivo., Conclusions: A high-quality two-dimensional and 3D in vivo visualization of the retinal structures and en face visualization of the retina and choroidal vascular plexus of vertebrates was possible. Our studies affirm that SS-OCT and SS-OCTA are viable methods for evaluating the in vivo retinal and choroidal structure across terrestrial, aquatic, and aerial vertebrates., Translational Relevance: In vivo characterization of retinal morphology and vasculature plexus of multiple species using SS-OCT and SS-OCTA imaging can increase the pool of species available as models of human retinal diseases.

Clustered regularly interspaced short palindromic repeats (CRISPR)-based genomic disruption of vascular endothelial growth factor A ( Vegfa ) with a single gRNA suppresses choroidal neovascularization (CNV) in preclinical studies, offering the prospect of long-term anti-angiogenesis therapy for neovascular age-related macular degeneration (AMD). Genome editing using CRISPR-CRISPR-associated endonucleases (Cas9) with multiple guide RNAs (gRNAs) can enhance gene-ablation efficacy by augmenting insertion-deletion (indel) mutations with gene truncations but may also increase the risk of off-target effects. In this study, we compare the effectiveness of adeno-associated virus (AAV)-mediated CRISPR-Cas9 systems using single versus paired gRNAs to target two different loci in the Vegfa gene that are conserved in human, rhesus macaque, and mouse. Paired gRNAs increased Vegfa gene-ablation rates in human cells in vitro but did not enhance VEGF suppression in mouse eyes in vivo . Genome editing using paired gRNAs also showed a similar degree of CNV suppression compared with single-gRNA systems. Unbiased genome-wide analysis using genome-wide unbiased identification of double-stranded breaks (DSBs) enabled by sequencing (GUIDE-seq) revealed weak off-target activity arising from the second gRNA. These findings suggest that in vivo CRISPR-Cas9 genome editing using two gRNAs may increase gene ablation but also the potential risk of off-target mutations, while the functional benefit of targeting an additional locus in the Vegfa gene as treatment for neovascular retinal conditions is unclear., Competing Interests: G.Y. received research support from Clearside Biomedical, Genentech, and Iridex and personal fees for consultancy from Abbvie, Adverum, Alimera, Anlong, Bausch & Lomb, Clearside Biomedical, Endogena, Genentech, Gyroscope Therapeutics, Intergalactic Therapeutics, Iridex, NGM Biopharmaceutical, Regeneron, Thea, Topcon, and Zeiss, all of which are unrelated to the contents of this study. The other authors have no relevant conflicts of interest., (© 2022 The Author(s).)

The visualization and assessment of retinal microvasculature are important in the study, diagnosis, monitoring, and guidance of treatment of ocular and systemic diseases. With the introduction of optical coherence tomography angiography (OCTA), it has become possible to visualize the retinal microvasculature volumetrically and without a contrast agent. Many lab-based and commercial clinical instruments, imaging protocols and data analysis methods and metrics, have been applied, often inconsistently, resulting in a confusing picture that represents a major barrier to progress in applying OCTA to reduce the burden of disease. Open data and software sharing, and cross-comparison and pooling of data from different studies are rare. These inabilities have impeded building the large databases of annotated OCTA images of healthy and diseased retinas that are necessary to study and define characteristics of specific conditions. This paper addresses the steps needed to standardize OCTA imaging of the human retina to address these limitations. Through review of the OCTA literature, we identify issues and inconsistencies and propose minimum standards for imaging protocols, data analysis methods, metrics, reporting of findings, and clinical practice and, where this is not possible, we identify areas that require further investigation. We hope that this paper will encourage the unification of imaging protocols in OCTA, promote transparency in the process of data collection, analysis, and reporting, and facilitate increasing the impact of OCTA on retinal healthcare delivery and life science investigations., (© 2022. The Author(s).)

Optoretinogram, a technique in which optical coherence tomography (OCT) is used to measure retinal functions in response to a visible light stimulus, can be a potentially useful tool to quantify retinal health alterations. Existing experimental studies on animals have focused on measuring the global retinal response by transversally averaging 3D data across the retina, which minimizes the spatial resolution of the signals, and limits the signal-to-noise ratio because only central B-scans are collected and analyzed. These problems were addressed in this study by collecting volumetric data to probe functional signals and developing an improved 3D registration approach to align such series-acquired OCT volumes. These data were then divided into small blocks and subject to a spatiotemporal analysis, whose results confirmed the spatial-dependence of functional signals. By further averaging, the overall measurement accuracies for the position and the scattering signals were estimated to be approximately 30 nm and 1.1 %, respectively. With improved accuracy, this method revealed certain novel functional signals that have not been previously reported. In conclusion, this work provides a powerful tool to monitor retinal local and global functional changes in aging, diseased, or treated rodent eyes., (© 2021 Wiley-VCH GmbH.)

Several specialized retinal optical coherence tomography (OCT) acquisition and processing methods have been recently developed to allow in vivo probing of light-evoked photoreceptors function, focusing on measurements in individual photoreceptors (rods and cones). Recent OCT investigations in humans and experimental animals have shown that the outer segments in dark-adapted rods and cones elongate in response to the visible optical stimuli that bleach fractions of their visual photopigment. We have previously successfully contributed to these developments by implementing OCT intensity-based "optoretinograms" (ORG), the paradigm of using near-infrared OCT (NIR OCT) to measure bleaching-induced back-scattering and/or elongation changes of photoreceptors in the eye in vivo . In parallel, several groups have successfully implemented phase-based ORGs, mainly in human studies, exploiting changes in the phases of back-scattered light. This allowed more sensitive observations of tiny alterations of photoreceptors structures. Applications of the phase-based ORG have been implemented primarily in high speed and cellular resolution AO-OCT systems that can visualize photoreceptor mosaic, allowing phase measurements of path length changes in outer segments of individual photoreceptors. The phase-based ORG in standard resolution OCT systems is much more demanding to implement and has not been explored extensively. This manuscript describes our efforts to implement a phase analysis framework to retinal images acquired with a standard resolution and raster scanning OCT system, which offers much lower phase stability than line-field or full-field OCT detection schemes due to the relatively slower acquisition speed. Our initial results showcase the successful extraction of phase-based ORG signal from the B-scans acquired at ∼100 Hz rate and its favorable comparison with intensity-based ORG signal extracted from the same data sets. We implemented the calculation of phase-based ORG signals using Knox-Thompson paths and modified signal recovery by adding decorrelation weights. The phase-sensitive ORG signal analysis developed here for mouse retinal raster scanning OCT systems could be in principle extended to clinical retinal raster scanning OCT systems, potentially opening doors for clinically friendly ORG probing., Competing Interests: The authors declare that there are no conflicts of interest related to this article., (© 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreement.)