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Purpose To quantify retinal capillary density and morphology in uveitis using spectral-domain optical coherence tomography angiography (SD-OCTA). Design Cross-sectional, observational study. Methods Healthy and uveitic subjects were recruited from 2 tertiary care eye centers. Prototype SD-OCTA devices (Cirrus; Carl Zeiss Meditec, Inc, Dublin, California, USA) were used to generate 3 × 3-mm 2 OCTA images centered on the fovea. Subjects were placed into 3 groups based on the type of optical microangiography (OMAG) algorithm used for image processing (intensity and/or phase) and type of retinal segmentation (automatic or manual). A semi-automated method was used to calculate skeleton density (SD), vessel density (VD), fractal dimension (FD), and vessel diameter index (VDI). Retinal vasculature was assessed in the superficial retinal layer (SRL), deep retinal layer (DRL), and nonsegmented retinal layer (NS-RL). A generalized estimating equations model was used to analyze associations between the OCTA measures and disease status within each retinal layer. A P value Results The SD, VD, and FD of the parafoveal capillaries were lower in uveitic eyes compared with healthy eyes in all retinal segments. In addition, SD and VD were significantly lower in the DRL of subjects with uveitic macular edema. There was no correlation in any capillary parameters and anatomic classification of uveitis. Conclusions Quantitative analysis of parafoveal capillary density and morphology in uveitis demonstrates significantly lower capillary density and complexity. SD-OCTA algorithms are robust enough to detect these changes and can provide a novel diagnostic index of disease for uveitis subjects.

An ultra high resolution spectral-domain optical coherence tomography (SD-OCT) together with an advanced animal restraint and positioning system was built for noninvasive non-contact in vivo three-dimensional imaging of rodent models of ocular diseases. The animal positioning system allowed the operator to rapidly locate and switch the areas of interest on the retina. This function together with the capability of precise spatial registration provided by the generated OCT fundus image allows the system to locate and compare the same lesion (retinal tumor in the current study) at different time point throughout the entire course of the disease progression. An algorithm for fully automatic segmentation of the tumor boundaries and calculation of tumor volume was developed. The system and algorithm were successfully applied to monitoring retinal tumor growth quantitatively over time in the LHBETATAG mouse model of retinoblastoma.

Rhodopsin, the light-sensing molecule in the outer segments of rod photoreceptors, is responsible for converting light into neuronal signals in a process known as phototransduction. Rhodopsin is thus a functional biomarker for rod photoreceptors. Here we report a novel technology based on visible-light optical coherence tomography (VIS-OCT) for in vivo molecular imaging of rhodopsin. The depth resolution of OCT allows the visualization of the location where the change of optical absorption occurs and provides a potentially accurate assessment of rhodopsin content by segmentation of the image at the location. Rhodopsin OCT can be used to quantitatively image rhodopsin distribution and thus assess the distribution of functional rod photoreceptors in the retina. Rhodopsin OCT can bring significant impact into ophthalmic clinics by providing a tool for the diagnosis and severity assessment of a variety of retinal conditions.

We developed an optical coherence photoacoustic microscopy (OC-PAM) system, which can accomplish optical coherence tomography (OCT) and photoacoustic microscopy (PAM) simultaneously by using a single pulsed broadband light source. With a center wavelength of 800 nm and a bandwidth of 30 nm, the system is suitable for imaging the retina. Generated from the same group of photons, the OCT and PAM images are intrinsically registered in the lateral directions. To test the capabilities of the system on multimodal ophthalmic imaging, we imaged the retina of pigmented rats. The OCT images showed the retinal structures with quality similar to conventional OCT, while the PAM images revealed the distribution of absorbers in the retina. Since the absorption of hemoglobin is relatively weak at around 800 nm, the NIR PAM signals are generated mainly from melanin in the posterior segment of the eye, thus providing melanin-specific imaging of the retina.

Glaucoma is a leading cause of irreversible blindness, presently affecting approximately 70 million people worldwide.1,2 The most common form is primary open-angle glaucoma (POAG), which accounts for approximately 90% of all cases. Ocular hypertension (OH) is the major risk factor for POAG and lowering IOP is the only effective treatment.3 However, many patients remain refractory to existing IOP-lowering interventions and eventually may become blind, underscoring the unmet medical need for novel approaches to control IOP. Intraocular pressure is a product of the rate of aqueous humor production, resistance to outflow, and episcleral venous pressure.3,4 The aqueous outflow pathways in the angle of the eye are composed of a spongy tissue called the trabecular meshwork (TBM), which leads into Schlemm's canal. Much of the resistance to outflow resides within the TBM, putatively within 7 to 14 μm of the inner wall of Schlemm's canal in a region known as the juxtacanalicular or cribriform region.5–,8 Primary open-angle glaucoma is caused by an increase in outflow resistance9,10 due to poorly understood defects in this region.11–,14 Ocular hypertension in POAG has been associated with dysregulated intracellular signaling pathways controlled by IL-1/NF-κB,15 TGF-β superfamily,16 and Wnt.17,18 The pathophysiology of OH leading to POAG shares many features with a secondary form of OH caused by treatment with glucocorticoids (GCs).19–,25 Glucocorticoids are a class of steroid hormones released in response to stress. As part of the natural feedback mechanism that turns down the inflammatory response, they are useful for treating a wide variety of diseases.26 Complicating this, however, is the considerable interindividual variability and tissue-specific GC sensitivity among individuals, which can cause a variety of systemic side effects.27–,31 Treatment with steroids, such as dexamethasone or triamcinolone acetonide (TA), in the eye causes elevated IOP in predisposed individuals. It has been documented that approximately 40% of the normal population develops an IOP increase > 6 mm Hg above baseline following topical administration of GCs, 4 times a day for 4 to 6 weeks.32 When GCs are administered intravitreally, IOP may increase by 30% or more in up to half of patients.33 These individuals are considered to be “steroid responders.” Interestingly, almost all POAG patients are steroid responders25,32,34–,40; conversely, steroid-responders who do not have POAG are at much higher risk of developing POAG compared to nonresponders.37–,41 A better understanding of genetic and molecular mechanisms underlying individual variation in the response to steroids could shed light on our understanding of steroid-induced OH and OH leading to POAG. That being said, mechanisms of steroid-induced OH and OH leading to POAG also show distinct differences; therefore, studying the genetics of steroid-induced OH specifically could lead to discovery of genes involved in IOP regulation that could not have been revealed any other way. The genome-wide association study (GWAS) represents an agnostic approach for prioritization of genes that are associated with a disease process, and makes it possible to discover novel genes that could not otherwise be identified. In the current study, we performed a pharmaco-GWAS for this purpose, using a cohort enrolled from patients treated with off-label intravitreal TA (IVTA) for various retinal indications, and followed up with an independent candidate gene study. We identified two independent quantitative trait loci (QTLs) at chromosomal locus 6p21.33 associated with the novel mucin gene HCG22.

Purpose To quantitatively evaluate the retinal microvasculature in human subjects with retinal venous occlusions (RVO) using optical coherence tomography angiography (OCTA). Design Retrospective, cross-sectional, observational case series. Participants Sixty subjects (84 eyes) were included (20 BRVO, 14 CRVO, 24 unaffected fellow eyes, and 26 controls). Methods OCTA was performed on a prototype, spectral domain-OCTA system in the 3x3mm central macular region. Custom software was used to quantify morphology and density of retinal capillaries using four quantitative parameters. The vasculature of the segmented retinal layers and nonsegmented whole retina were analyzed. Main outcome measures Fractal dimension (FD), vessel density (VD), skeletal density (SD), and vessel diameter index (VDI) within the segmented retinal layers and nonsegmented whole retina vasculature. Results Nonsegmented analysis of RVO eyes demonstrated significantly lower FD (1.64±0.01 vs 1.715±0.002; p

To analyze glaucomatous eyes with known focal defects of the nerve fiber layer (NFL), relating optical coherence tomography (OCT) findings to clinical examination, NFL and stereoscopic optic nerve head (ONH) photography, and Humphrey 24-2 visual fields.Cross-sectional prevalence study.The authors followed 19 patients in the study group and 14 patients in the control group.Imaging with OCT was performed circumferentially around the ONH with a circle diameter of 3.4 mm using an internal fixation technique. One hundred OCT scan points taken within 2.5 seconds were analyzed.Measurements of NFL thickness using OCT were performed.In most eyes with focal NFL defects, OCTs showed significant thinning of the NFL in areas closely corresponding to focal defects visible on clinical examination, to red-free photographs, and to defects on the Humphrey visual fields. Optical coherence tomography enabled the detection of focal defects in the NFL with a sensitivity of 65% and a specificity of 81%.Analysis of NFL thickness in eyes with focal defects showed good structural and functional correlation with clinical parameters. Optical coherence tomography contributes to the identification of focal defects in the NFL that occur in early stages of glaucoma.

Objective: This study aimed to develop a protocol to screen and monitor patients with diabetic macular thickening using optical coherence tomography (OCT), a technique for high-resolution cross-sectional imaging of the retina. Design: A cross-sectional pilot study was conducted. Participants: A total of 182 eyes of 107 patients with diabetic retinopathy, 55 eyes from 31 patients with diabetes but no ophthalmoscopic evidence of retinopathy, and 73 eyes from 41 healthy volunteers were studied. Intervention: Six optical coherence tomograms were obtained in a radial spoke pattern centered on the fovea. Retinal thickness was computed automatically from each tomogram at a total of 600 locations throughout the macula. Macular thickness was displayed geographically as a false-color topographic map and was reported numerically as averages in each of nine regions. Main Outcome Measures: Correlation of OCT with slit-lamp biomicroscopy, fluorescein angiography, and visual acuity was measured. Results: Optical coherence tomography was able to quantify the development and resolution of both foveal and extrafoveal macular thickening. The mean ± standard deviation foveal thickness was 174 ± 18 μ m in normal eyes, 179 ± 17 μ m in diabetic eyes without retinopathy, and 256 ± 114 μ m in eyes with nonproliferative diabetic retinopathy. Foveal thickness was highly correlated among left and right eyes of normal eyes (mean ± standard deviation difference of 6 ± 9 μ m). Foveal thickness measured by OCT correlated with visual acuity ( r 2 = 0.79). A single diabetic eye with no slit-lamp evidence of retinopathy showed abnormal foveal thickening on OCT. Conclusions: Optical coherence tomography was a useful technique for quantifying macular thickness in patients with diabetic macular edema. The topographic mapping protocol provided geographic information on macular thickness that was intuitive and objective.

Purpose: Optical coherence tomography (OCT) is a new technology that uses near-infrared light in an interferometer to produce approximately 10-µm resolution crosssectional images of the tissue of interest. The authors performed repeated quantitative assessment of nerve fiber layer thickness in individuals with normal and glaucomatous eyes, and they evaluated the reproducibility of these measurements. Methods: The authors studied 21 eyes of 21 subjects by OCT. Each subject underwent five repetitions of a series of scans on five separate occasions within a 1-month period. Each series consisted of three circular scans around the optic nerve head (diameters, 2.9, 3.4, and 4.5 mm). Each series was performed separately using internal (fixation with same eye being studied) and external (fixation with contralateral eye) fixation techniques. The eye studied and the sequence of testing were assigned randomly. Results: Internal fixation (IF), in general, provides a slightly higher degree of reproducibility than external fixation (EF). Reproducibility was better in a given eye on a given visit than from visit to visit. Reproducibility as measured by intraclass correlation coefficients were as follows: circle diameter (CD), 2.9 mm, 0.51/0.57 (normal/glaucoma) (IF), 0.43/0.54 (EF); CD, 3.4 mm, 0.56/0.52 (IF), 0.43/0.61 (EF); CD, 4.5 mm, 0.53/0.43 (IF), 0.42/0.49 (EF). Conclusions: Nerve fiber layer thickness can be reproducibly measured using OCT. Internal is superior to external fixation; each circle diameter tested provides adequate reproducibility.

Purpose: The authors describe the clinical feature of ten patients with a new syndrome characterized by the presence of retinal vasculitis, multiple macroaneurysms, neuro-retinitis, and peripheral capillary nonperfusion. Methods: The authors evaluated ten patients identified to have clinical features compatible with the syndrome of idiopathic retinal vasculitis, aneurysms and neuroretinitis (IRVAN). Clinical examination findings, sequential fundus photographs (when available), fluorescein angiograms, systemic investigations, response to therapy, and visual outcomes were reviewed. Results: Seven eyes of four patients sustained a marked decrease in visual acuity of 20/200 or worse. Visual loss was due to a combination of an exudative maculopathy and sequelae of retinal ischemia. Capillary nonperfusion was seen in all ten patients and was severe enough to warrant panretinal laser photocoagulation in six patients. Systemic investigations were uniformly noncontributory. Oral prednisone appears to have little beneficial effects on patients with this disorder. Conclusions: Patients with IRVAN have characteristic retinal features that readily identify this syndrome. An increased awareness of this rare syndrome may help to identify sight-threatening complications at an earlier stage. The authors caution against extensive medical investigations. Ophthalmology 1995;102:1089-1097

Purpose To quantify changes in retinal microvasculature in diabetic retinopathy (DR) by using spectral-domain optical coherence tomography angiography (SD-OCTA). Methods Retrospective, cross-sectional, observational study of healthy and diabetic adult subjects with and without DR. Retinal microvascular changes were assessed by using SD-OCTA images and an intensity-based optical microangiography algorithm. A semiautomated program was used to calculate indices of microvascular density and morphology in nonsegmented and segmented SD-OCTA images. Microvascular density was quantified by using skeleton density (SD) and vessel density (VD), while vessel morphology was quantified as fractal dimension (FD) and vessel diameter index (VDI). Statistical analyses were performed by using the Student's t-test or analysis of variance with post hoc Tukey honest significant difference tests for multiple comparisons. Results Eighty-four eyes with DR and 14 healthy eyes were studied. Spearman's rank test demonstrated a negative correlation between DR severity and SD, VD, and FD, and a positive correlation with VDI (ρ = −0.767, −0.7166, −0.768, and +0.5051, respectively; P < 0.0001). All parameters showed high reproducibility between graders (ICC = 0.971, 0.962, 0.937, and 0.994 for SD, VD, FD, and VDI, respectively). Repeatability (κ) was greater than 0.99 for SD, VD, FD, and VDI. Conclusions Vascular changes in DR can be objectively and reliably characterized with SD, VD, FD, and VDI. In general, decreasing capillary density (SD and VD), branching complexity (FD), and increasing average vascular caliber (VDI) were associated with worsening DR. Changes in capillary density and morphology were significantly correlated with diabetic macular edema.

We combined photoacoustic ophthalmoscopy (PAOM) with autofluorescence imaging for simultaneous in vivo imaging of dual molecular contrasts in the retina using a single light source. The dual molecular contrasts come from melanin and lipofuscin in the retinal pigment epithelium (RPE). Melanin and lipofuscin are two types of pigments and are believed to play opposite roles (protective versus exacerbate) in the RPE in the aging process. We have successfully imaged the retina of pigmented and albino rats at different ages. The experimental results showed that multimodal PAOM system can be a potentially powerful tool in the study of age-related degenerative retinal diseases.

To assess the rate of infectious endophthalmitis and to describe the clinical and microbiological features of eyes that develop clinically suspected endophthalmitis after an intravitreal injection of vascular endothelial growth factor antagonists.The medical records of patients undergoing intravitreal injections of anti-vascular endothelial growth factor agents from January 1, 2005, through December 31, 2010, at a single university referral center and associated satellite clinics were retrospectively analyzed to determine the rate of infectious endophthalmitis after intravitreal anti-vascular endothelial growth factor injections.Twelve cases (11 patients) of clinically suspected endophthalmitis were identified after a total of 60,322 injections (0.02%; 95% confidence interval, 0.0114%-0.0348%). Of the 12 cases, 11 presented within 3 days of the injection. Of the 7 culture-positive cases, 5 were because of Streptococcus species. In 4 of the 5 Streptococcus cases, final visual acuity was hand motions or worse. The rate of clinically suspected endophthalmitis was 0.018% after bevacizumab and 0.027% after ranibizumab injections.A very low rate of endophthalmitis after intravitreal injections of anti-vascular endothelial growth factor agents was observed. Patients typically presented within 3 days of injection. Streptococcus species was the most common bacteria isolated, and it was generally associated with poor visual outcomes.

Photoacoustic ophthalmoscopy (PAOM) is a new retinal imaging technology that offers the unique capability to measure optical absorption in the retina. Because PAOM is compatible with optical coherence tomography, scanning laser ophthalmoscopy, and autofluorescence imaging, registered multimodal images can be acquired from a single device at comparable resolution for comprehensive anatomic and functional retinal characterizations. Therefore, PAOM is anticipated to have applications in both research and clinical diagnosis of many blinding diseases. The authors explain the basic principles of the photoacoustic effect and imaging. Then, different types of photoacoustic microscopy are introduced and compared. Finally, the current status of photoacoustic imaging in animal eyes is presented and the prospects of future development of PAOM are suggested.

We have developed a non-invasive photoacoustic ophthalmoscopy (PAOM) for in vivo retinal imaging. PAOM detects the photoacoustic signal induced by pulsed laser light shined onto the retina. By using a stationary ultrasonic transducer in contact with the eyelids and scanning only the laser light across the retina, PAOM provides volumetric imaging of the retinal micro-vasculature and retinal pigment epithelium at a high speed. For B-scan frames containing 256 A-lines, the current PAOM has a frame rate of 93 Hz, which is comparable with state-of-the-art commercial spectral-domain optical coherence tomography (SD-OCT). By integrating PAOM with SD-OCT, we further achieved OCT-guided PAOM, which can provide multi-modal retinal imaging simultaneously. The capabilities of this novel technology were demonstrated by imaging both the microanatomy and microvasculature of the rat retina in vivo.

Segmentation of retinal layers from OCT images is fundamental to diagnose the progress of retinal diseases. In this study we show that the retinal layers can be automatically and/or interactively located with good accuracy with the aid of local coherence information of the retinal structure. OCT images are processed using the ideas of texture analysis by means of the structure tensor combined with complex diffusion filtering. Experimental results indicate that our proposed novel approach has good performance in speckle noise removal, enhancement and segmentation of the various cellular layers of the retina using the STRATUSOCTTM system.

A high-speed spectral-domain optical coherence tomography (OCT) system was built to image the human retina in vivo. A fundus image similar to the intensity image produced by a scanning laser ophthalmoscope (SLO) was generated from the same spectra that were used for generating the OCT sectional images immediately after the spectra were collected. This function offers perfect spatial registration between the sectional OCT images and the fundus image, which is desired in ophthalmology for monitoring data quality, locating pathology, and increasing reproducibility. This function also offers a practical way to detect eye movements that occur during the acquisition of the OCT image. The system was successfully applied to imaging human retina in vivo.

We determine the reliability and reproducibility of retinal thickness measurements with a custom-built OCT retinal image analy- sis software OCTRIMA. Ten eyes of five healthy subjects undergo repeated standard macular thickness map scan sessions by two expe- rienced examiners using a Stratus OCT device. Automatic/semi auto- matic thickness quantification of the macula and intraretinal layers is performed using OCTRIMA software. Intraobserver, interobserver, and intervisit repeatability and reproducibility coefficients, and intraclass correlation coefficients ICCs per scan are calculated. Intraobserver, interobserver, and intervisit variability combined account for less than 5% of total variability for the total retinal thickness measurements and less than 7% for the intraretinal layers except the outer segment/ reti- nal pigment epithelium RPE junction. There is no significant differ- ence between scans acquired by different observers or during different visits. The ICCs obtained for the intraobserver and intervisit variability tests are greater than 0.75 for the total retina and all intraretinal layers, except the inner nuclear layer intraobserver and interobserver test and the outer plexiform layer, intraobserver, interobserver, and intervisit test. Our results indicate that thickness measurements for the total retina and all intraretinal layers except the outer segment/RPE junc- tion performed using OCTRIMA are highly repeatable and reproducible. © 2009 Society of Photo-Optical Instrumentation Engineers. DOI: 10.1117/1.3268773

To compare ultrahigh-resolution optical coherence tomography (UHR-OCT) technology to a standard-resolution OCT instrument for the imaging of macular hole pathology and repair; to identify situations where UHR-OCT provides additional information on disease morphology, pathogenesis, and management; and to use UHR-OCT as a baseline for improving the interpretation of the standard-resolution images.Observational and interventional case series.Twenty-nine eyes of 24 patients clinically diagnosed with macular hole in at least one eye.A UHR-OCT system has been developed and employed in a tertiary-care ophthalmology clinic. Using a femtosecond laser as the low-coherence light source, this new UHR-OCT system can achieve an unprecedented 3-mum axial resolution for retinal OCT imaging. Comparative imaging was performed with UHR-OCT and standard 10-mum resolution OCT in 29 eyes of 24 patients with various stages of macular holes. Imaging was also performed on a subset of the population before and after macular hole surgery.Ultrahigh- and standard-resolution cross-sectional OCT images of macular hole pathologies.Both UHR-OCT and standard-resolution OCT exhibited comparable performance in differentiating various stages of macular holes. The UHR-OCT provided improved imaging of finer intraretinal structures, such as the external limiting membrane and photoreceptor inner segment (IS) and outer segment (OS), and identification of the anatomy of successful surgical repair. The improved resolution of UHR-OCT enabled imaging of previously unidentified changes in photoreceptor morphology associated with macular hole pathology and postoperative repair. Visualization of the junction between the photoreceptor IS and OS was found to be an important indicator of photoreceptor integrity for both standard-resolution and UHR-OCT images.Ultrahigh-resolution optical coherence tomography improves the visualization of the macular hole architectural morphology. The increased resolution of UHR-OCT enables the visualization of photoreceptor morphology associated with macular holes. This promises to lead to a better understanding of the pathogenesis of macular holes, the causes of visual loss secondary to macular holes, the timing of surgical repair, and the evaluation of postsurgical outcome. Ultrahigh-resolution optical coherence tomography imaging of macular holes that correspond to known alterations in retinal morphology can be used to interpret retinal morphology in UHR-OCT images. Comparisons of UHR-OCT images with standard-resolution OCT images can establish a baseline for the better interpretation of clinical standard-resolution OCT images. The ability to visualize photoreceptors and their integrity or impairment is an indicator of macular hole progression and surgical outcome.

Keeping the air quality acceptable has become an important task for decision makers as well as for non-governmental organizations. Particulate and gaseous emissions of pollutant from industries and auto-exhausts are responsible for rising discomfort, increasing airway diseases, decreasing productivity and the deterioration of artistic and cultural patrimony in urban centers. A model to determine the air quality in urban areas using a geographical information system will be presented here. This system permits the integration, handling, analysis and simulation of spatial and temporal data of the ambient concentration of the main pollutant. It allows the users to characterize and recognize areas with a potential increase or improvement in its air pollution situation. It is also possible to compute past or present conditions by changing basic input information as traffic flow, or stack emission rates. Additionally the model may be used to test the compliance of local standard air quality, to study the environmental impact of new industries or to determine the changes in the conditions when the vehicle circulation is increased.

Purpose To test the capability of a novel dual-beam Doppler optical coherence tomography (OCT) technique for simultaneous in vivo measurement of the Doppler angle and, thus, the absolute retinal blood velocity and the retinal flow rate, without the influence of motion artifacts. Methods A novel dual-beam Doppler spectral domain OCT (SD-OCT) was developed. The two probing beams are separated with a controllable distance along an arbitrary direction, both of which are controlled by two independent 2D optical scanners. Two sets of optical Doppler tomography (ODT) images are acquired simultaneously. The Doppler angle of each blood vessel segment is calculated from the relative coordinates of the centers of the blood vessel in the two corresponding ODT images. The absolute blood flow velocity and the volumetric blood flow rate can then be calculated. To measure the total retinal blood flow, we used a circular scan pattern centered at the optic disc to obtain two sets of concentric OCT/ODT images simultaneously. Results We imaged two normal human subjects at ages of 48 and 34 years. The total retinal blood flow rates of the two human subjects were calculated to be 47.01 μL/min (older subject) and 51.37 μL/min (younger subject), respectively. Results showed that the performance of this imaging system is immune to eye movement, since the two sets of ODT images were acquired simultaneously. Conclusions The dual-beam OCT/ODT system is successful in measuring the absolute retinal blood velocity and the volumetric flow rate. The advantage of the technique is that the two sets of ODT images used for the calculation are acquired simultaneously, which eliminates the influence of eye motion and ensures the accuracy of the calculated hemodynamic parameters.

The ultimate goal of the study is to provide an imaging tool to detect the earliest signs of glaucoma before clinically visible damage occurs to the retinal nerve fiber layer (RNFL). Studies have shown that the optical reflectance of the damaged RNFL at short wavelength (560 nm) is reduced much more than that at long wavelength, which provides spectral contrast for imaging the earliest damage to the RNFL. To image the spectral contrast we built a dual-band spectral-domain optical coherence tomography (SD-OCT) centered at 808 nm (NIR) and 415 nm (VIS). The light at the two bands was provided by the fundamental and frequency-doubled outputs of a broadband Ti:Sapphire laser. The depth resolution of the NIR and VIS OCT systems are 4.7 µm and 12.2 µm in the air, respectively. The system was applied to imaging the rat retina in vivo. Significantly different appearances between the OCT cross sectional images at the two bands were observed. The ratio of the light reflected from the RNFL over that reflected from the entire retina at the two bands were quantitatively compared. The experimental results showed that the dual-band OCT system is feasible for imaging the spectral contrasts of the RNFL.

We investigated the feasibility of simultaneously imaging two distinctive molecular contrasts provided by the absorbed photons in biological tissues with a single light source. The molecular contrasts are based on two physical effects induced by the absorbed photons: photoacoustics (PA) and autofluorescence (AF). In an integrated multimodal imaging system, the PA and AF signals were detected by a high-sensitivity ultrasonic transducer and an avalanche photodetector, respectively. The system was tested by imaging ocular tissue samples, including the retinal pigment epithelium and the ciliary body. The acquired images provided information on the spatial distributions of melanin and lipofuscin in these samples.

We have developed an adaptive optics photoacoustic microscope (AO-PAM) for high-resolution imaging of biological tissues, especially the retina. To demonstrate the feasibility of AO-PAM we first designed the AO system to correct the wavefront errors of the illuminating light of PAM. The aberrations of the optical system delivering the illuminating light to the sample in PAM was corrected with a close-loop AO system consisting of a 141-element MEMS-based deformable mirror (DM) and a Shack-Hartmann (SH) wavefront sensor operating at 15 Hz. The photoacoustic signal induced by the illuminating laser beam was detected by a custom-built needle ultrasonic transducer. When the wavefront errors were corrected by the AO system, the lateral resolution of PAM was measured to be better than 2.5 µm using a low NA objective lens. We tested the system on imaging ex vivo ocular samples, e.g., the ciliary body and retinal pigment epithelium (RPE) of a pig eye. The AO-PAM images showed significant quality improvement. For the first time we were able to resolve single RPE cells with PAM.

Since the photoacoustic effect relies only on the absorbed optical energy, the back-reflected photons from samples in optical-resolution photoacoustic microscopy are usually discarded. By employing a 2 x 2 single-mode fiber optical coupler in a laser-scanning optical-resolution photoacoustic microscope for delivering the illuminating laser light and collecting the back reflected photons, a fiber-optic confocal microscope is integrated with the photoacoustic microscope. Thus, simultaneous multimodal imaging can be achieved with a single light source and images from the two modalities are intrinsically registered. Such capabilities are demonstrated in imaging both phantoms and small animals in vivo.

We have successfully imaged the retinal tumor in a mouse model using an ultra-high resolution spectral-domain optical coherence tomography (SD-OCT) designed for small animal retinal imaging. For segmentation of the tumor boundaries and calculation of the tumor volume, we developed a novel segmentation algorithm. The algorithm is based on parametric deformable models (active contours) and is driven by machine learning-based region classification, namely a Conditional Random Field. With this algorithm we are able to obtain the tumor boundaries automatically, while the user can specify additional constraints (points on the boundary) to correct the segmentation result, if needed. The system and algorithm were successfully applied to studies on retinal tumor progression and monitoring treatment effects quantitatively in a mouse model of retinoblastoma.

Purpose To demonstrate the application of high-resolution spectral-domain optical coherence tomography (SD-OCT) for three-dimensional (3D) retinal imaging of small animals and quantitative retinal information extraction using 3D segmentation of the OCT images. Methods A high-resolution SD-OCT system was built for in vivo imaging of rodent retina. OCT fundus images similar to those acquired with a scanning laser ophthalmoscope (SLO) were constructed from the measured OCT data, which provided precise spatial registration of the OCT cross-sectional images on the fundus. A 3D segmentation algorithm was developed for calculation of the retinal thickness map. OCT images were compared by histologic examination. Results High-quality OCT images of the retinas of mice (B6/SJLF2 for normal retina, rhodopsin-deficient Rho(-/-) for photoreceptor degeneration, and LH(BETA)T(AG) for retinoblastoma) and rat (Wistar) were acquired. The OCT images compared well with histology. Not only was a 3D image of the tumor in a retinoblastoma mouse model successfully imaged in vivo but the tumor volume was extracted from the 3D image. Retinal thickness maps were calculated that enabled successful quantitative comparison of the retinal thickness distribution between the normal (202.3 +/- 9.3 microm) and the degenerative (102.7 +/- 12.6 microm) mouse retina. Conclusions High-resolution spectral-domain OCT provides unprecedented high-quality 2D and 3D in vivo visualization of retinal structures of mouse and rat models of retinal diseases. With the capability of 3D quantitative information extraction and precise spatial registration, the OCT system made possible longitudinal study of ocular diseases that has been impossible to conduct.

Optical Doppler tomography (ODT) is a branch of optical coherence tomography (OCT) that can measure the speed of a blood flow by measuring the Doppler shift impinged on the probing sample light by the moving blood cells. However, the measured speed of blood flow is a function of the Doppler angle, which needs to be determined in order to calculate the absolute velocity of the blood flow inside a vessel. We developed a technique that can extract the Doppler angle from the 3D data measured with spectral-domain OCT, which needs to extract the lateral and depth coordinates of a vessel in each measured ODT and OCT image. The lateral coordinates and the diameter of a blood vessel were first extracted in each OCT structural image by using the technique of blood vessel shadowgram, a technique first developed by us for enhancing the retinal blood vessel contrast in the en face view of the 3D OCT. The depth coordinate of a vessel was then determined by using a circular averaging filter moving in the depth direction along the axis passing through the vessel center in the ODT image. The Doppler angle was then calculated from the extracted coordinates of the blood vessel. The technique was applied in blood flow measurements in retinal blood vessels, which has potential impact on the study and diagnosis of blinding diseases like glaucoma.

The development of improved segmentation algorithms for more consistently accurate detection of retinal boundaries is a potentially useful solution to the limitations of existing optical coherence tomography (OCT) software. We modeled artifacts related to operator errors that may normally occur during OCT imaging and evaluated their influence on segmentation results using a novel segmentation algorithm. These artifacts included: defocusing, depolarization, decentration, and a combination of defocusing and depolarization. Mean relative reflectance and average thickness of the automatically extracted intraretinal layers was then measured. Our results show that defocusing and depolarization errors together have the greatest altering effect on all measurements and on segmentation accuracy. A marked decrease in mean relative reflectance and average thickness was observed due to depolarization artifact in all intraretinal layers, while defocus resulted in a less-marked decrease. Decentration resulted in a marked but not significant change in average thickness. Our study demonstrates that care must be taken for good-quality imaging when measurements of intraretinal layers using the novel algorithm are planned in future studies. An awareness of these pitfalls and their possible solutions is crucial for obtaining a better quantitative analysis of clinically relevant features of retinal pathology.

We previously developed a technique to acquire a SLO (scanning laser ophthalmoscope) like fundus intensity image from the raw spectra measured with spectral-domain optical coherence tomography (OCT), the same spectra used to generate a 3D OCT data set. This technique offers simultaneous fundus and OCT images and, therefore, solves the problem of registering a cross sectional OCT image to fundus features. However, the registration of high density OCT images is still an unsolved problem because no useful fundus image can be generated from the high density scans. High density OCT images can significantly improve the image quality and enhance the visualization of retinal structure, especially the structure of small lesions. We have developed a feature-based algorithm, which can register a high density OCT image on the fundus image generated from normal density scans. The algorithm was successfully tested for both normal and diseased eyes.

To determine the rate of anatomical closure of idiopathic macular holes undergoing vitreous surgery with respect to preoperative horizontal diameter as determined by optical coherence tomography (OCT) and to correlate postoperative visual acuity, duration of symptoms, and late reopening with initial idiopathic macular hole diameter by OCT.Forty eyes of 40 patients with an idiopathic macular hole were examined with OCT before and after vitreous surgery. All eyes were treated with pars plana vitrectomy, peeling of posterior cortical vitreous, and dilute perfluoropropane or sulfur hexafluoride gas. Face-down positioning was maintained for 7 to 14 days.Twenty-two (92%) of 24 eyes with a preoperative idiopathic macular hole diameter smaller than 400 microm measured by OCT attained anatomical closure following surgery. Anatomical closure was observed in 9 (56%) of 16 eyes with a macular hole diameter of 400 microm or larger measured by OCT (P =.02). The median postoperative visual acuity improvement was 4 Snellen lines in the 31 eyes achieving anatomical closure and no change in the 9 eyes not achieving anatomical closure (P.001). Late macular hole reopening at longer than 6 months occurred in 3 (10%) of 31 eyes with an initially closed macular hole. This event was observed only in macular holes 400 microm or larger measured by OCT. The preoperative macular hole diameter (P =.02) and duration of symptoms (P =.02) were factors predictive of anatomical closure of the macular hole postoperatively.The postoperative closure of idiopathic macular holes following vitreous surgery was related to the preoperative macular hole diameter determined by OCT, with lesions smaller than 400 microm demonstrating higher success rates. A trend toward greater visual acuity improvement was demonstrated for idiopathic macular holes smaller than 400 microm. Late reopening was only seen in macular holes that were 400 microm or larger measured by OCT. Preoperative analysis and measurement of idiopathic macular holes with OCT may help delineate postoperative expectations for successful anatomical closure of the macular hole, visual acuity, and long-term closure.

We report the first study of laser-tissue interaction in the femtosecond time regime. Retinal damage thresholds and mechanisms produced by exposure to high-intensity femtosecond laser pulses were investigated in chinchilla grey rabbits. Exposures were performed using single laser pulses of 80 fs duration at 625 nm. ED 50 injury thresholds of 0.75 and 4.5 μJ were measured using fluorescein angiographic and ophthalmoscopic visibility criteria evaluating 204 laser exposures. Ultrastructural studies including light and electron microscopy were performed on selected lesions. Results suggest that the primary energy deposition in the retina occurs in melanin, However, in contrast to laser injuries produced by longer pulses, exposures of more than 100 × threshold in the 50-100 \mu J range did not produce significantly more severe lesions or hemorrhage. This suggests the presence of a nonlinear damage limiting mechanics in tissue exposed to femtosecond laser pulses.

Q -switched and mode-locked neodymium: YAG laser pulses can be used to produce optical breakdown inside the eye, permitting surgical disruption of transparent or pigmented ocular tissues. The physical processes responsible for the initiation, propagation, and control of this important and novel class of laser tissue interactions are reviewed.