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Potentially blinding corneal diseases alter the morphology of the human cornea. At the early stages of disease development, these changes occur at the cellular level. The ability to visualize and quantify such changes can lead to early diagnostics of corneal pathologies, which is pivotal for the long-term preservation of vision. Here we present a Powell Lens-based Line-Field Optical Coherence Tomography system that combines high spatial resolution (2.4 μm × 2.2 μm × 1.7 μm (x × y × z)) in biological tissue, sufficient to resolve individual cells, high sensitivity (90.5 dB), sufficient to image the semi-transparent human cornea, and fast image acquisition rate (~ 2,400 fps), sufficient to suppress most involuntary eye motion artifacts and allow for contactless, in-vivo imaging of the cellular structure of the human cornea. Volumetric images acquired in-vivo from corneas of healthy subjects show epithelial, endothelial and keratocytes cells, as well as sub-basal and stromal nerves. The system's high axial resolution also allows for volumetric morphometry of the corneal endothelium, Descemet's membrane and the pre-Descemet's (Dua) layer., (© 2024. The Author(s).)

The trade-off between transverse resolution and depth-of-focus (DOF) typical for optical coherence tomography (OCT) systems based on conventional optics, prevents "single-shot" acquisition of volumetric OCT images with sustained high transverse resolution over the entire imaging depth. Computational approaches for correcting defocus and higher order aberrations in OCT images developed in the past require highly stable phase data, which poses a significant technological challenge. Here, we present an alternative computational approach to sharpening OCT images and reducing speckle noise, based on intensity OCT data. The novel algorithm uses non-local priors to model correlated speckle noise within a maximum a posteriori framework to generate sharp and noise-free images. The performance of the algorithm was tested on images of plant tissue (cucumber) and in-vivo healthy human cornea, acquired with line-field spectral domain OCT (LF-SD-OCT) systems. The novel algorithm effectively suppressed speckle noise and sharpened or recovered morphological features in the OCT images for depths up to 13×DOF (depth-of-focus) relative to the focal plane., Competing Interests: The authors declare no conflicts of interest related to this article., (© 2024 Optica Publishing Group.)

Dynamic optical coherence tomography (dOCT) utilizes time-dependent signal intensity fluctuations to enhance contrast in OCT images and indirectly probe physiological processes in cells. Majority of the dOCT studies published so far are based on acquisition of 2D images (B-scans or C-scans) by utilizing point-scanning Fourier domain (spectral or swept-source) OCT or full-field OCT respectively, primarily due to limitations in the image acquisition rate. Here we introduce a novel, high-speed spectral domain line-field dOCT (SD-LF-dOCT) system and image acquisition protocols designed for fast, volumetric dOCT imaging of biological tissues. The imaging probe is based on an exchangeable afocal lens pair that enables selection of combinations of transverse resolution (from 1.1 µm to 6.4 µm) and FOV (from 250 × 250 µm 2 to 1.4 × 1.4 mm 2 ), suitable for different biomedical applications. The system offers axial resolution of ∼ 1.9 µm in biological tissue, assuming an average refractive index of 1.38. Maximum sensitivity of 90.5 dB is achieved for 3.5 mW optical imaging power at the tissue surface and maximum camera acquisition rate of 2,000 fps. Volumetric dOCT images acquired with the SD-LF-dOCT system from plant tissue (cucumber), animal tissue (mouse liver) and human prostate carcinoma spheroids allow for volumetric visualization of the tissues' cellular and sub-cellular structures and assessment of cellular motility., Competing Interests: The authors declare no conflicts of interest., (© 2024 Optica Publishing Group.)

Digital correction of optical aberrations allows for high-resolution imaging across the full depth range in optical coherence tomography (OCT). Many digital aberration correction (DAC) methods have been proposed in the past to evaluate and correct monochromatic error in OCT images. However, other factors that deteriorate the image quality have not been fully investigated. Specifically, in a broadband line-scan spectral-domain OCT system (LS-SD-OCT), photons with different wavelengths scattered from the same transverse location and in the imaged object will be projected onto different spatial coordinates onto the 2D camera sensor, which in this work is defined as spatial-spectral crosstalk. In addition, chromatic aberrations in both axial and lateral directions are not negligible for broad spectral bandwidths. Here we present a novel approach to digital recovery of the spatial resolution in images acquired with a broadband LS-SD-OCT, which addresses these two main factors that limit the effectiveness of DAC for restoring diffraction-limited resolution in LS-SD-OCT images. In the proposed approach, spatial-spectral crosstalk and chromatic aberrations are suppressed by the registration of monochromatic sub-band tomograms that are digitally corrected for aberrations. The new method was validated by imaging a standard resolution target, a microspheres phantom, and different biological tissues. LS-SD-OCT technology combined with the proposed novel image reconstruction method could be a valuable research tool for various biomedical and clinical applications., Competing Interests: The authors declare no conflicts of interest related to this article., (© 2023 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

A Powell lens is used in a line-field spectral domain OCT (PL-LF-SD-OCT) system to generate a line-shaped imaging beam with almost uniform distribution of the optical power in the line direction. This design overcomes the severe sensitivity loss (∼10 dB) observed along the line length direction (B-scan) in LF-OCT systems based on cylindrical lens line generators. The PL-LF-SD-OCT system offers almost isotropic spatial resolution (Δx and Δy ∼2 µm, Δz ∼1.8 µm) in free space and sensitivity of ∼87 dB for 2.5 mW imaging power at 2,000 fps imaging rate with only ∼1.6 dB sensitivity loss along the line length. Images acquired with the PL-LF-SD-OCT system allow for visualization of the cellular and sub-cellular structure of biological tissues., Competing Interests: The authors declare no conflicts of interest related to this article., (© 2023 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

A novel decorrelation-based approach for measuring localized transverse flow velocity using line-scan (LS) optical coherence tomography (OCT) is proposed. The new approach allows for separation of the flow velocity component along the line-illumination direction of the imaging beam from other orthogonal velocity components, from particle diffusion motion, and from noise-induced distortion in the OCT signal's temporal autocorrelation. The new method was verified by imaging flow in a glass capillary and a microfluidic device and mapping the spatial distribution of the flow velocity within the beam's illumination plane. This method can be extended in the future to map the three-dimensional flow velocity fields for both ex -vivo and in-vivo applications., Competing Interests: The authors declare no conflicts of interest related to this article., (© 2023 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

In-vivo , non-contact, volumetric imaging of the cellular and sub-cellular structure of the human cornea and limbus with optical coherence tomography (OCT) is challenging due to involuntary eye motion that introduces both motion artifacts and blur in the OCT images. Here we present the design of a line-scanning (LS) spectral-domain (SD) optical coherence tomography system that combines 2 × 3 × 1.7 µm (x, y, z) resolution in biological tissue with an image acquisition rate of ∼2,500 fps, and demonstrate its ability to image in-vivo and without contact with the tissue surface, the cellular structure of the human anterior segment tissues. Volumetric LS-SD-OCT images acquired over a field-of-view (FOV) of 0.7 mm × 1.4 mm reveal fine morphological details in the healthy human cornea, such as epithelial and endothelial cells, sub-basal nerves, as well as the cellular structure of the limbal crypts, the palisades of Vogt (POVs) and the blood microvasculature of the human limbus. LS-SD-OCT is a promising technology that can assist ophthalmologists with the early diagnostics and optimal treatment planning of ocular diseases affecting the human anterior eye., Competing Interests: The authors declare no conflicts of interest related to this article., (© 2022 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

Purpose: The purpose of this study was to characterize the central epithelial thickness (CET) of penetrating keratoplasty corneal specimens obtained from patients with keratoconus (KC) and correlate the histological patterns with their clinical history., Methods: Ex vivo histological imaging was performed to measure CET and total corneal thickness (TCT) in 56 patients with KC. Microscopic slides from penetrating keratoplasty corneal specimens, stained with hematoxylin and eosin were evaluated using bright field microscopy. CET and TCT were measured, and morphological features were studied. Clinical history regarding duration of KC prior to surgery and length of and tolerance to contact lens wear were compared and analyzed., Results: The microscopic slides of all patients available for follow up (n=48) were analyzed and CET and TCT were measured. The histological evaluation revealed 3 distinctive epithelial patterns. Pattern 1 with central hypertrophic and hydropic changes (n=19) measured 70.89±25.88μm in CET and 308.63±100.74μm in TCT; Pattern 2 (n=14) had not changed, similar to normal epithelium CET and TCT measuring 36.5±7.02μm and 260.14±87.93μm respectively. Pattern 3 (n=15) demonstrated thinner central epithelium characterized by atrophy and focal hydropic changes measuring 19.93±4.60μm and 268.00±79.39μm in CET and TCT respectively (all p<0.0001). The presence of Pattern 2 characterized by similar to normal CET was correlated with the duration of the condition (R=0.600, p=0.030). There was a significant difference in the length of CL wear comparing those with patterns 1 and 2 versus 3 (least no. of CL years) (p=0.05 and p=0.33 respectivelly)., Conclusions: Patients with advanced disease have various central corneal epithelial changes detected with histology. Although each central epithelial pattern type was distinctive comparing the 3 patterns, there was no correlation with years of CL wear but only with the duration of the condition., (Copyright © 2020 Spanish General Council of Optometry. Published by Elsevier España, S.L.U. All rights reserved.)

We present the first spectral domain optical coherence tomography (SD-OCT) system that combines an isotropic imaging resolution of ~1.5 µm in biological tissue with a 250 kHz image acquisition rate, for in vivo non-contact, volumetric imaging of the cellular structure of the human cornea. OCT images of the healthy human cornea acquired with this system reveal the cellular structure of the corneal epithelium, cellular debris and mucin clusters in the tear film, the shape, size and spatial distribution of the sub-basal corneal nerves and keratocytes in the corneal stroma, as well as reflections from endothelial nuclei. The corneal images presented here demonstrate the potential clinical value of the new high speed, high resolution OCT system for non-invasive diagnostics and monitoring the treatment of corneal diseases., Competing Interests: The authors declare that there are no conflicts of interest related to this article.

Morphological and functional changes in the rat retina and optic nerve head (ONH), associated with 8 weeks of intermittent moderately elevated intraocular pressure (IOP) were measured with a combined ultrahigh resolution optical coherence tomography (UHR-OCT) and electroretinography (ERG) system. The IOP of male Sprague-Dawley rats was raised in one eye to ~35 mmHg for 1 hour/day on 6 days each week using vascular loops. Single-flash ERG traces and volumetric UHR-OCT images of the ONH were acquired from both eyes before, during and after IOP elevations at weeks 1, 5 and 9 of the study. The UHR-OCT images showed depression of the posterior eye around the ONH during the IOP elevations, the magnitude of which increased significantly from week 1 to week 9 (p = 0.01). The ERG a-wave and b-wave amplitudes increased temporarily during IOP elevations and returned to normal ~30 minutes after loop removal. Recurrent intermittent IOP spikes caused > 30% decrease in the ERG a-wave and b-wave amplitudes measured during the IOP elevations over the course of 2 months. This study suggests that recurrent, relatively short-duration IOP spikes for extended period of time are associated with peri-ONH tissue hypercompliance and reduced retinal functional response to visual stimulation during acute IOP elevation.

A novel image processing algorithm based on a modified Bayesian residual transform (MBRT) was developed for the enhancement of morphological and vascular features in optical coherence tomography (OCT) and OCT angiography (OCTA) images. The MBRT algorithm decomposes the original OCT image into multiple residual images, where each image presents information at a unique scale. Scale selective residual adaptation is used subsequently to enhance morphological features of interest, such as blood vessels and tissue layers, and to suppress irrelevant image features such as noise and motion artefacts. The performance of the proposed MBRT algorithm was tested on a series of cross-sectional and enface OCT and OCTA images of retina and brain tissue that were acquired in-vivo . Results show that the MBRT reduces speckle noise and motion-related imaging artefacts locally, thus improving significantly the contrast and visibility of morphological features in the OCT and OCTA images., Competing Interests: The authors declare that there are no conflicts of interest related to this article.

Acute elevation of intraocular pressure (IOP) to ischemic and non-ischemic levels can cause temporary or permanent changes in the retinal morphology, function and blood flow/blood perfusion. Previously, such changes in the retina were assessed separately with different methods in clinical studies and animal models. In this study, we used a combined OCT+ ERG system in combination with Doppler OCT and OCT angiography (OCTA) imaging protocols, in order to evaluate simultaneously and correlate changes in the retinal morphology, the retinal functional response to visual stimulation, and the retinal blood flow/blood perfusion, associated with IOP elevation to ischemic and non-ischemic levels in rats. Results from this study suggest that the inner retina responds faster to IOP elevation to levels greater than 30 mmHg with significant reduction of the total retinal blood flow (TRBF), decrease of the capillaries' perfusion and reduction of the ON bipolar cells contribution to the ERG traces. Furthermore, this study showed that ischemic levels of IOP elevation cause an additional significant decrease in the ERG photoreceptor response in the posterior retina. Thirty minutes after IOP normalization, retinal morphology, blood flow and blood perfusion recovered to baseline values, while retinal function did not recover completely.

Purpose: To establish a relationship between the diameter and depth position of vessels in the superior and inferior corneo-scleral limbus using ultra-high resolution optical coherence tomography (UHR-OCT)., Methods: Volumetric OCT images of the superior and inferior limbus were acquired from 14 healthy subjects with a research-grade UHR-OCT system. Differences in vessel diameter and depth between superior and inferior limbus were analyzed using repeated measured ANOVA in SPSS and R., Results: The mean (± SD) superior and inferior diameters were 29±18μm and 24±18μm respectively, and the mean (± SD) superior and inferior depths were 177±109μm and 207±132μm respectively. The superior limbal vessels were larger than the inferior ones (RM-ANOVA, p=0.004), and the inferior limbal vessels were deeper than the superior vessels (RM-ANOVA, p=0.041). There was a positive linear association between limbal vessel depth and size within the superior and inferior limbus with Pearson correlation coefficients of 0.803 and 0.754, respectively., Conclusion: This study demonstrated that the UHR-OCT was capable of imaging morphometric characteristics such as the size and depth of vessels in the limbus. The results of this study suggest a difference in the size and depth of vessels across different positions of the limbus, which may be indicative of adaptations to chronic hypoxia caused by the covering of the superior limbus by the upper eyelid. UHR-OCT may be a useful tool to evaluate the effect of contact lenses on the microvascular properties within the limbus., (Copyright © 2017 Spanish General Council of Optometry. All rights reserved.)

The variability in the spatial orientation of retinal blood vessels near the optic nerve head (ONH) results in imprecision of the measured Doppler angle and therefore the pulsatile blood flow (BF), when those parameters are evaluated using Doppler OCT imaging protocols based on dual-concentric circular scans. Here, we utilized a dense concentric circle scanning protocol and evaluated its precision for measuring pulsatile retinal BF in rats for different numbers of the circular scans. An spectral domain optical coherence tomography (SD-OCT) system operating in the 1060-nm spectral range with image acquisition rate of 47,000 A-scans/s was used to acquire concentric circular scans centered at the rat's ONH, with diameters ranging from 0.8 to 1.0 mm. A custom, automatic blood vessel segmentation algorithm was used to track the spatial orientation of the retinal blood vessels in three dimensions, evaluate the spatially dependent Doppler angle and calculate more accurately the axial BF for each major retinal blood vessel. Metrics such as retinal BF, pulsatility index, and resistance index were evaluated for each and all of the major retinal blood vessels. The performance of the proposed dense concentric circle scanning protocols was compared with that of the dual-circle scanning protocol. Results showed a 3.8±2.2  deg difference in the Doppler angle calculation between the two approaches, which resulted in ∼7% difference in the calculated retinal BF., ((2017) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).)

A research-grade OCT system was used to image in-vivo and without contact with the tissue, the cellular structure and microvasculature of the healthy human corneo-scleral limbus. The OCT system provided 0.95 µm axial and 4 µm (2 µm) lateral resolution in biological tissue depending on the magnification of the imaging objective. Cross-sectional OCT images acquired tangentially from the inferior limbus showed reflective, loop-like features that correspond to the fibrous folds of the palisades of Vogt (POV). The high OCT resolution allowed for visualization of individual cells inside the limbal crypts, capillaries extending from the inside of the POV's fibrous folds and connecting to a lateral grid of micro-vessels located in the connective tissue directly below the POV, as well as reflections from individual red blood cells inside the capillaries. Difference in the reflective properties of the POV was observed among subjects of various pigmentation levels of the POV. Morphological features observed in the high resolution OCT images correlated well with histology. The ability to visualize the limbal morphology and microvasculature in-vivo at cellular level can aid the diagnostics and treatment of limbal stem cell dysfunction and dystrophies.

Corneal degenerative conditions such as keratoconus (KC) cause progressive damage to the anterior corneal tissue and eventually severely compromise visual acuity. The ability to visualize corneal tissue damage in-vivo at cellular or sub-cellular level at different stages of development of KC and other corneal diseases, can aid the early diagnostics as well as the development of more effective treatment approaches for various corneal pathologies, including keratoconus. Here, we present the optical design of an optical coherence tomography system that can achieve 0.95 µm axial resolution in biological tissue and provide test results for the system's spatial resolution and sensitivity. Corneal images acquired in-vivo with this system from healthy and keratoconic human subjects reveal the cellular and sub-cellular structure of the corneal epithelium, as well as the normal and abnormal structure of the Bowman's membrane and the anterior corneal stroma.

The probability density function (PDF) of light scattering intensity can be used to characterize the scattering medium. We have recently shown that in optical coherence tomography (OCT), a PDF formalism can be sensitive to the number of scatterers in the probed scattering volume and can be represented by the K-distribution, a functional descriptor for non-Gaussian scattering statistics. Expanding on this initial finding, here we examine polystyrene microsphere phantoms with different sphere sizes and concentrations, and also human skin and fingernail in vivo. It is demonstrated that the K-distribution offers an accurate representation for the measured OCT PDFs. The behavior of the shape parameter of K-distribution that best fits the OCT scattering results is investigated in detail, and the applicability of this methodology for biological tissue characterization is demonstrated and discussed.

Purpose: To visualize in vivo and quantify the thickness of the posterior corneal layers: the acellular pre-Descemet's layer (PDL), Descemet's membrane (DM), and endothelium (END) in healthy subjects, using ultrahigh-resolution optical coherence tomography (UHR-OCT)., Methods: A research-grade, 800-nm UHR-OCT system with 0.95-μm axial resolution in corneal tissue was used to image in vivo the posterior cornea in healthy subjects. The system offers approximately 98 dB sensitivity for 680 μW optical power incident on the cornea and 34,000 A-scans/s image acquisition rate. This study comprised 20 healthy subjects, aged 20 to 60 years. The thickness of the PDL, DM, and END layers was measured both with a custom, automatic segmentation algorithm and manually., Results: The boundaries and structure of the posterior corneal layers were clearly visible in the UHR-OCT images. The average thickness was measured to be 6.6 ± 1.4 μm (PDL), 10.4 ± 2.9 μm (DM), and 4.8 ± 0.4 μm (END), which agrees well with published data from ex vivo studies. Both the END and DM thickness showed minor spatial variations, whereas the PDL showed up to 2× thickness change for different locations on the same cross-sectional corneal image or over the entire imaged region of the cornea., Conclusions: Our data indicate that all three layers of the posterior cornea can be clearly visualized in vivo and their thicknesses measured precisely with UHR-OCT. Although the PDL thickness showed large spatial variations, the thickness of the DM and END layers was consistent over the entire imaged region of the cornea.

Purpose: Moderately elevated intraocular pressure (IOP) is a risk factor for open-angle glaucoma. Some patients suffer glaucoma despite clinically measured normal IOPs. Fluctuations in IOP may have a significant role since IOPs are higher during sleep and inversion activities. Controlled transient elevations of IOPs in rats over time lead to optic nerve structural changes that are similar to the early changes observed in constant chronic models of glaucoma. Because early intervention decreases glaucoma progression, this study was done to determine if early physiological changes to the retina could be detected with noninvasive electrophysiological and optical imaging tests during moderately elevated IOP., Methods: Intraocular pressures were raised to moderately high levels (35 mm Hg) in one eye of Sprague-Dawley rats while the other (control) eye was untreated. One group of rats underwent scotopic threshold response (STR) and electroretinogram (ERG) testing, while another 3 groups underwent optical coherence tomography (OCT) imaging, Western blot, or histologic evaluation., Results: The amplitudes of the STR and ERG responses in eyes with moderately elevated IOPs were enhanced compared to the values before IOP elevation, and compared to untreated contralateral eyes. Structural changes to the optic nerve also occurred during IOP elevation., Conclusions: Although ischemic IOP elevations are well-known to globally reduce components of the scotopic ERG, acute elevation in rats to levels often observed in untreated glaucoma patients caused an increase in these parameters. Further exploration of these phenomena may be helpful in better understanding the mechanisms mediating early retinal changes during fluctuating or chronically elevated IOP.

Background: Optical coherence tomography (OCT) is a minimally invasive imaging technique, which utilizes the spatial and temporal coherence properties of optical waves backscattered from biological material. Recent advances in tunable lasers and infrared camera technologies have enabled an increase in the OCT imaging speed by a factor of more than 100, which is important for retinal imaging where we wish to study fast physiological processes in the biological tissue. However, the high scanning rate causes proportional decrease of the detector exposure time, resulting in a reduction of the system signal-to-noise ratio (SNR). One approach to improving the image quality of OCT tomograms acquired at high speed is to compensate for the noise component in the images without compromising the sharpness of the image details., Methods: In this study, we propose a novel reconstruction method for rapid OCT image acquisitions, based on a noise-compensated homotopic modified James-Stein non-local regularized optimization strategy. The performance of the algorithm was tested on a series of high resolution OCT images of the human retina acquired at different imaging rates., Results: Quantitative analysis was used to evaluate the performance of the algorithm using two state-of-art denoising strategies. Results demonstrate significant SNR improvements when using our proposed approach when compared to other approaches., Conclusions: A new reconstruction method based on a noise-compensated homotopic modified James-Stein non-local regularized optimization strategy was developed for the purpose of improving the quality of rapid OCT image acquisitions. Preliminary results show the proposed method shows considerable promise as a tool to improve the visualization and analysis of biological material using OCT.

Improving the spatial resolution of Optical Coherence Tomography (OCT) images is important for the visualization and analysis of small morphological features in biological tissue such as blood vessels, membranes, cellular layers, etc. In this paper, we propose a novel reconstruction approach to obtaining super-resolved OCT tomograms from multiple lower resolution images. The proposed Multi-Penalty Conditional Random Field (MPCRF) method combines four different penalty factors (spatial proximity, first and second order intensity variations, as well as a spline-based smoothness of fit) into the prior model within a Maximum A Posteriori (MAP) estimation framework. Test carried out in retinal OCT images illustrate the effectiveness of the proposed MPCRF reconstruction approach in terms of spatial resolution enhancement, as compared to previously published super resolved image reconstruction methods. Visual assessment of the MPCRF results demonstrate the potential of this method in better preservation of fine details and structures of the imaged sample, as well as retaining the sharpness of biological tissue boundaries while reducing the effects of speckle noise inherent to OCT. Quantitative evaluation using imaging metrics such as Signal-to-Noise Ratio (SNR), Contrast to Noise Ratio (CNR), Equivalent Number of Looks (ENL), and Edge Preservation Parameter show significant visual quality improvement with the MPCRF approach. Therefore, the proposed MPCRF reconstruction approach is an effective tool for enhancing the spatial resolution of OCT images without the necessity for significant imaging hardware modifications.

Optical coherence tomography (OCT) allows for non-invasive 3D visualization of biological tissue at cellular level resolution. Often hindered by speckle noise, the visualization of important biological tissue details in OCT that can aid disease diagnosis can be improved by speckle noise compensation. A challenge with handling speckle noise is its inherent non-stationary nature, where the underlying noise characteristics vary with the spatial location. In this study, an innovative speckle noise compensation method is presented for handling the non-stationary traits of speckle noise in OCT imagery. The proposed approach centers on a non-stationary spline-based speckle noise modeling strategy to characterize the speckle noise. The novel method was applied to ultra high-resolution OCT (UHROCT) images of the human retina and corneo-scleral limbus acquired in-vivo that vary in tissue structure and optical properties. Test results showed improved performance of the proposed novel algorithm compared to a number of previously published speckle noise compensation approaches in terms of higher signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR) and better overall visual assessment.

An optical coherence tomography (OCT) microvascular imaging platform, consisting of Doppler (DOCT) and speckle variance (svOCT) modalities, and microvascular image quantification tools are developed. The quantification methods extract blood flow-related parameters from DOCT images and vessel morphological parameters from svOCT images. This platform is used to assess the microvascular (DOCT and svOCT) images obtained during a clinical study on late oral radiation toxicity. This specific pathology was considered a suitable scenario for verifying the performance of the developed quantification platform because late oral radiation toxicity is known to involve microvascular damage. The derived parameters are compared between several DOCT and svOCT images from one patient and one healthy volunteer as proof-of-principle, and the significance of the observed differences is discussed. Given the low number of OCT clinical studies that measure and quantify microvascular images and considering the importance of such quantification in a number of pathologies, this newly developed platform can serve as a useful tool in studying diseases and treatments with microvascular involvement.

An ultrahigh resolution spectral domain optical coherence tomography (SD-OCT) system is used to observe for the first time in vivo the early effect of sodium iodate (NaIO3) toxicity on retinal morphology. Retinal degeneration is induced in rats via tail vein injection of NaIO3 and structural changes in the outer retina are assessed longitudinally at baseline and 1, 2, 3, 6, 8, and 10 h, and 12 post drug administration with OCT, H&E histology, and IgG immunochemistry. Disruption of the structural integrity and changes in the optical reflectivity of the photoreceptor inner (IS) and outer segment (OS) layers are observed as early as 1 h post NaIO3 injection. A new layer is observed in the OCT tomograms to form between the retinal pigmented epithelium and the photoreceptors OS a few hours post NaIO3 injection. The dynamics and the low optical reflectivity of this layer, as well as cell swelling and disruption of the blood-retina barrier observed in the histological and immunohistochemistry cross-sections suggest that the layer corresponds to temporary fluid accumulation in the retina. Results from this study demonstrate the effectiveness of OCT technology for monitoring dynamic changes in the retinal morphology and provide better understanding of the early stages of outer retina degeneration induced by NaIO3 toxicity.

High quality, large size volumetric imaging of biological tissue with optical coherence tomography (OCT) requires large number and high density of scans, which results in large data acquisition volume. This may lead to corruption of the data with motion artifacts related to natural motion of biological tissue, and could potentially cause conflicts with the maximum permissible exposure of biological tissue to optical radiation. Therefore, OCT can benefit greatly from different approaches to sparse or compressive sampling of the data where the signal is recovered from its sub-Nyquist measurements. In this paper, a new energy-guided compressive sensing approach is proposed for improving the quality of images acquired with Fourier domain OCT (FD-OCT) and reconstructed from sparse data sets. The proposed algorithm learns an optimized sampling probability density function based on the energy distribution of the training data set, which is then used for sparse sampling instead of the commonly used uniformly random sampling. It was demonstrated that the proposed energy-guided learning approach to compressive FD-OCT of retina images requires 45% fewer samples in comparison with the conventional uniform compressive sensing (CS) approach while achieving similar reconstruction performance. This novel approach to sparse sampling has the potential to significantly reduce data acquisition while maintaining image quality.

Purpose: To demonstrate the ability of high speed, ultrahigh-resolution optical coherence tomography (UHR-OCT) to measure and characterize in vivo visual stimulus-specific pupil dynamics in birds., Methods: Ten two-week old White Leghorn (Gallus gallus domesticus) chickens were imaged in this study. The chickens were dark-adapted for 1 hour and anesthetized with 2% isoflurane prior to the imaging procedure. Blue, green, and red single flash visual stimuli of 7 ms duration were used to evoke pupillary responses. UHR-OCT cross-sectional images of the pupil were acquired prior, during, and for several seconds after the visual stimuli onset. Images were processed with a novel custom automatic algorithm, designed to determine the pupil diameter changes over time., Results: Results from this study show that the pupillary constriction begins with the onset of the visual stimuli; however, maximum pupil constriction occurs ∼150 ms later. No statistically significant variation in the timing of the maximum pupillary constriction was observed for stimuli of different colors. However, significant variation was observed in the maximum pupil constriction amplitudes, between red-green and red-blue stimuli, but not between blue-green stimuli. Furthermore, the magnitude of the maximum pupil constriction decreased monotonically with time under isoflurane anesthesia., Conclusions: We demonstrated, for the first time, measurements of visually evoked pupillary dynamics in animals using high speed UHR-OCT. The results suggest dependence of the pupillary dynamics on the color of the visual stimulus, and adverse effects of isoflurane anesthesia on the visually evoked pupillary responses in chickens.

A spectral domain optical coherence tomography (SD-OCT) system and an oral imaging probe have been developed to visualize the microstructural morphology and microvasculature in the human oral cavity. Structural OCT images of ex vivo pig oral tissues with the histology of the same sites were acquired and compared for correlations. Structural in vivo OCT images of healthy human tissue as well as a pathologic site (ulcer) were obtained and analyzed based on the results of the ex vivo pig study, drawing on the similarity between human and swine oral tissues. In vivo Doppler and speckle variance OCT images of the oral cavity in human volunteers were also acquired, to demonstrate the feasibility of microvascular imaging of healthy and pathologic (scar) oral tissue.

The resolution in optical coherence tomography imaging is an important parameter which determines the size of the smallest features that can be visualized. Sparse sampling approaches have shown considerable promise in producing high resolution OCT images with fewer camera pixels, reducing both the cost and the complexity of an imaging system. In this paper, we propose a non-local approach to the reconstruction of high resolution OCT images from sparsely sampled measurements. An iterative strategy is introduced for minimizing a homotopic, non-local regularized functional in the spatial domain, subject to data fidelity constraints in the k-space domain. The novel algorithm was tested on human retinal, corneal, and limbus images, acquired in-vivo, demonstrating the effectiveness of the proposed approach in generating high resolution reconstructions from a limited number of camera pixels., (© 2012 Optical Society of America)

Purpose: To provide in vivo quantitative assessment of sodium iodate-induced retinal damage in a rat model of outer retinal degeneration using ultrahigh resolution optical coherence tomography (UHR-OCT)., Methods: Outer retinal degeneration was induced in four female Long Evans rats via tail vein injection of sodium iodate (40 mg/kg). Changes in the thickness and optical reflectivity of individual retinal layers were extracted using a semi-automatic segmentation algorithm and were assessed in vivo at 6 hours, days 1, 3, and 7, and up to 3 months post injection with UHR-OCT. Hematoxylin and eosin (H&E) histology was used to confirm the morphologic changes observed in the UHR-OCT images., Results: UHR-OCT tomograms showed progressive structural damage in the rat retina over time, such as swelling, thinning, complete disintegration of individual retinal layers, and clustering of highly reflective cellular debris. Photoreceptor swelling was observed 6 hours after injection of sodium iodate, followed by progressive structural decomposition of the outer retina. At 3 months post injection, the outer retina was completely disintegrated, and the inner nuclear layer (INL) was in direct contact with the choroid. Changes in the reflectivity of individual retinal layers were observed over time and correlated well with the morphologic changes., Conclusions: UHR-OCT permits in vivo, noninvasive, longitudinal, quantitative assessment of the progressive changes in retinal morphology and optical reflectivity in a sodium iodate rodent model of outer retinal degeneration.

We demonstrate the efficiency of the convolution using an optimized Kaiser-Bessel window to resample nonlinear data in wavenumber for Fourier-domain optical coherence tomography (OCT). We extend our previous experimental demonstration that was performed with a specific swept-source nonlinearity. The method is now applied to swept-source OCT data obtained for various simulated swept-source nonlinearities as well as spectral-domain OCT data obtained from both simulations and experiments. Results show that the new optimized method is the most efficient for handling all the different types of nonlinearities in the wavenumber domain that one can encounter in normal practice. The efficiency of the method is evaluated through comparison with common methods using resampling through interpolation prior to performing a fast-Fourier transform and with the accurate but time-consuming discrete Fourier transform for unequally spaced data, which involves Vandermonde matrices.

Visually evoked fast intrinsic optical signals (IOSs) were recorded for the first time in vivo from all layers of healthy chicken retina by using a combined functional optical coherence tomography (fOCT) and electroretinography (ERG) system. The fast IOSs were observed to develop within ∼5 ms from the on-set of the visual stimulus, whereas slow IOSs were measured up to 1 s later. The visually evoked IOSs and ERG traces were recorded simultaneously, and a clear correlation was observed between them. The ability to measure visually evoked fast IOSs non-invasively and in vivo from individual retinal layers could significantly improve the understanding of the complex communication between different retinal cell types in healthy and diseased retinas.

A novel optical coherence tomography (OCT) reconstruction approach is introduced for improved visualization of inner-retina capillaries in retinal OCT tomograms. The proposed method utilizes a minimization framework based on a tensor total variation (TTV) energy functional, to enforce capillary structural characteristics in the spatial domain. By accounting for structure tensor characteristics, the TTV reconstruction method allows for contrast enhancement of capillary structural characteristics. The novel TTV method was tested on high resolution OCT images acquired in-vivo from the foveal region of the retina of a healthy human subject. Experimental results demonstrate significant contrast and visibility enhancement of the inner retina capillaries in the retinal OCT tomograms, achieved by use of the TTV reconstruction method. Therefore, the TTV method has a strong potential for improved disease progression analysis based on the study of disease-induced changes in the inner retina vasculature., (2011 Optical Society of America)

Keratocytes are fibroblast-like cells that maintain the optical clarity and the overall health of the cornea. The ability to measure precisely their density and spatial distribution in the cornea is important for the understanding of corneal healing processes and the diagnostics of some corneal disorders. A novel computerized approach to detection and counting of keratocyte cells from ultra high resolution optical coherence tomography (UHR-OCT) images of the human corneal stroma is presented. The corneal OCT data is first processed using a state-of-the-art despeckling algorithm to reduce the effect of speckle on detection accuracy. A thresholding strategy is then employed to allow for improved delineation of keratocyte cells by suppressing similarly shaped features in the data, followed by a second-order moment analysis to identify potential cell nuclei candidates. Finally, a local extrema strategy is used to refine the candidates to determine the locations and the number of keratocyte cells. Cell density distribution analysis was carried in 3D UHR-OCT images of the human corneal stroma, acquired in-vivo. The cell density results obtained using the proposed novel approach correlate well with previous work on computerized keratocyte cell counting from confocal microscopy images of human cornea.

The limbus is the structurally rich transitional region of tissue between the cornea on one side, and the sclera and conjunctiva on the other. This zone, among other things, contains nerves passing to the cornea, blood and lymph vasculature for oxygen and nutrient delivery and for waste, CO(2) removal and drainage of the aqueous humour. In addition, the limbus contains stem cells responsible for the existence and healing of the corneal epithelium. Here we present 3D images of the healthy human limbus, acquired in vivo with a spectral domain optical coherence tomography system operating at 1060nm. Cross-sectional and volumetric images were acquired from temporal and nasal locations in the human limbus with ~3µm x 18µm (axial x lateral) resolution in biological tissue at the rate of 92,000 A-scans/s. The imaging enabled detailed mapping of the corneo-scleral tissue morphology, and visualization of structural details such as the Vogt palisades, the blood and lymph vasculature including the Schlemm's canal and the trabecular meshwork, as well as corneal nerve fiber bundles. Non-invasive, volumetric, high resolution imaging reveals fine details of the normal human limbal structure, and promises to provide invaluable information about its changes in health and disease as well as during and after corneal surgery.

The chicken retina is an established animal model for myopia and light-associated growth studies. It has a unique morphology: it is afoveate and avascular; oxygen and nutrition to the inner retina is delivered by a vascular tissue (pecten) that protrudes into the vitreous. Here we present, to the best of our knowledge, the first in vivo, volumetric high-resolution images of the chicken retina. Images were acquired with an ultrahigh-resolution optical coherence tomography (UHROCT) system with 3.5 µm axial resolution in the retina, at the rate of 47,000 A-scans/s. Spatial variations in the thickness of the nerve fiber and ganglion cell layers were mapped by segmenting and measuring the layer thickness with a semi-automatic segmentation algorithm. Volumetric visualization of the morphology and morphometric analysis of the chicken retina could aid significantly studies with chicken retinal models of ophthalmic diseases.

Purpose: To evaluate the change in thickness of the anterior, stromal, and posterior corneal laminae in response to hypoxia-induced corneal swelling, by means of ultrahigh-resolution optical coherence tomography (UHR-OCT)., Methods: A UHR-OCT system, operating in the 1060-nm range, was used to acquire in vivo cross-sectional images of human cornea with a 3.2x10-microm (axial x lateral) resolution in corneal tissue. Corneal edema was induced by inserting a thick, positive-powered, soft contact lens, over which the eye was closed and patched for 3 hours. Tomograms were acquired from eight non-contact-lens wearers. Baseline images were obtained before inducing corneal edema, immediately after removal of the patch and the lens, and then every 15 minutes for approximately 2 hours. All images were postprocessed with a segmentation algorithm to identify the laminae visible in the image. The apical thickness of the laminae (epithelium [EPI], epithelial-Bowman's membrane [Ep-BM] complex, stroma, and endothelial-Descemet's membrane [En-DM] complex) were determined at each time interval., Results: There was an interaction between time after removal of the hypoxic stimulus and deswelling of the layers (RM-ANOVA; P<0.001). The epithelial and stromal thickness reduced significantly with time (P=0.001; P<0.001, respectively), whereas the Ep-BM and En-DM complexes did not (P>0.50). All layers except the En-DM complex exhibited a biphasic pattern of recovery., Conclusions: UHR-OCT showed regional differences in swelling due to hypoxic provocation. On removal of the hypoxic stimulus, the rate of recovery varied between layers, and all layers except the En-DM complex exhibited a biphasic recovery.

An important image post-processing step for optical coherence tomography (OCT) images is speckle noise reduction. Noise in OCT images is multiplicative in nature and is difficult to suppress due to the fact that in addition the noise component, OCT speckle also carries structural information about the imaged object. To address this issue, a novel speckle noise reduction algorithm was developed. The algorithm projects the imaging data into the logarithmic space and a general Bayesian least squares estimate of the noise-free data is found using a conditional posterior sampling approach. The proposed algorithm was tested on a number of rodent (rat) retina images acquired in-vivo with an ultrahigh resolution OCT system. The performance of the algorithm was compared to that of the state-of-the-art algorithms currently available for speckle denoising, such as the adaptive median, maximum a posteriori (MAP) estimation, linear least squares estimation, anisotropic diffusion and wavelet-domain filtering methods. Experimental results show that the proposed approach is capable of achieving state-of-the-art performance when compared to the other tested methods in terms of signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), edge preservation, and equivalent number of looks (ENL) measures. Visual comparisons also show that the proposed approach provides effective speckle noise suppression while preserving the sharpness and improving the visibility of morphological details, such as tiny capillaries and thin layers in the rat retina OCT images.

A computational model was developed to evaluate the limitations to the highest axial resolution, achievable with ultrahigh resolution optical coherence tomography (UHROCT) in the 1060 nm wavelength region for in-vivo imaging of the human and rodent retina. The model considers parameters such as the wavelength dependent water absorption, the average length of the human and rodent eyes, and the power limitations for the imaging beam as defined in the ANSI standard. A custom-built light source with re-shaped spectrum was used to verify experimentally the results from the computational model. Axial OCT resolution of 4.2 microm and 7.7 microm was measured from a mirror reflection with the custom light source by propagating the imaging beam through water cells with 5 mm and 25 mm thickness, corresponding to the average axial length of the rodent and human eye respectively. Assuming an average refractive index of 1.38 for retinal tissue, the expected axial OCT resolution in the rodent and human retina is 3 microm and 5.7 microm respectively. Retinal tomograms acquired in-vivo from the rat eye with the modified light source show clear visualization of all intraretinal layers, as well as a network of capillaries (approximately 10 microm in diameter) in the inner- and outer plexiform layers of the retina.

Retinal layer thickness, evaluated as a function of spatial position from optical coherence tomography (OCT) images is an important diagnostics marker for many retinal diseases. However, due to factors such as speckle noise, low image contrast, irregularly shaped morphological features such as retinal detachments, macular holes, and drusen, accurate segmentation of individual retinal layers is difficult. To address this issue, a computer method for retinal layer segmentation from OCT images is presented. An efficient two-step kernel-based optimization scheme is employed to first identify the approximate locations of the individual layers, which are then refined to obtain accurate segmentation results for the individual layers. The performance of the algorithm was tested on a set of retinal images acquired in-vivo from healthy and diseased rodent models with a high speed, high resolution OCT system. Experimental results show that the proposed approach provides accurate segmentation for OCT images affected by speckle noise, even in sub-optimal conditions of low image contrast and presence of irregularly shaped structural features in the OCT images.

A novel speckle noise reduction algorithm based on a combination of Anisotropic Diffusion (AD) filtering and Interval Type-II fuzzy system was developed for reducing speckle noise in Optical Coherence Tomography (OCT) images. Unlike regular AD, the Interval Type-II fuzzy based AD algorithm considers the uncertainty in the calculated diffusion coefficient and appropriate adjustments to the coefficient are made. The new algorithm offers flexibility in optimizing the trade-off between the two image metrics: signal-to-noise (SNR) and Edginess, which are directly related to the structure of the imaged object. Application of the Interval Type-II fuzzy AD algorithm to OCT tomograms acquired in-vivo from a human finger tip and human retina show reduction in the speckle noise with very little edge blurring and about 13 dB and 7 dB image SNR improvement respectively. Comparison with Wiener, Adaptive Lee and regular Anisotropic Diffusion filters, applied to the same images, demonstrates the superior performance of the fuzzy Type-II AD algorithm in terms image SNR and edge preservation metrics improvement.

A novel speckle reduction technique based on soft thresholding of wavelet coefficients using interval type II fuzzy system was developed for reducing speckle noise in Optical Coherence Tomography images. The proposed algorithm is an extension of a recently published method for filtering additive Gaussian noise by use of type I fuzzy system. Unlike type I, interval type II fuzzy based thresholding filter considers the uncertainty in the calculated threshold and the wavelet coefficient is adjusted based on this uncertainty. A single parameter controls the signal-to-noise (SNR) improvement. Application of this novel algorithm to optical coherence tomograms acquired in-vivo from a human finger tip show reduction in the speckle noise with little edge blurring and image SNR improvement of about 10dB. Comparison with adaptive Wiener and adaptive Lee filters, applied to the same image, demonstrated the superior performance of the fuzzy type II algorithm in terms of image metrics improvement.

The ability of ultra-high-resolution optical coherence tomography (UHR OCT) to discriminate between healthy and pathological human brain tissue is examined by imaging ex vivo tissue morphology of various brain biopsies. Micrometer-scale OCT resolution (0.9x2 microm, axialxlateral) is achieved in biological tissue by interfacing a state-of-the-art Ti:Al2O3 laser (lambda(c)=800 nm, delta lambda=260 nm, and P(out)=120 mW exfiber) to a free-space OCT system utilizing dynamic focusing. UHR OCT images are acquired from both healthy brain tissue and various types of brain tumors including fibrous, athypical, and transitional meningioma and ganglioglioma. A comparison of the tomograms with standard hematoxylin and eosin (H&E) stained histological sections of the imaged biopsies demonstrates the ability of UHR OCT to visualize and identify morphological features such as microcalcifications (>20 microm), enlarged nuclei of tumor cells (approximately 8 to 15 microm), small cysts, and blood vessels, which are characteristic of neuropathologies and normally absent in healthy brain tissue., (Copyright 2005 Society of Photo-Optical Instrumentation Engineers)