[Show abstract][Hide abstract] ABSTRACT: The corneal endothelium (CE) is a single layer of cells lining the posterior face of the cornea providing metabolic functions essential for maintenance of corneal transparency. Adult CE cells lack regenerative potential and the number of CE cells decreases throughout life. In order to determine if endogenous DNA damage contributes to the age-related spontaneous loss of CE, we characterized CE in Ercc1(-/Δ) mice, which have impaired capacity to repair DNA damage and age prematurely. Eyes from 4.5-6 month-old Ercc1(-/Δ) mice, age-matched wild-type (WT) littermates, and old WT mice (24-34 months-old) were compared by spectral domain optical coherence tomography and corneal confocal microscopy. Histopathological changes in CE were further identified in paraffin tissue sections, whole mount immunostaining, and scanning- and transmission- electron microscopy. The CE of old WT mice displayed polymorphism and polymegathism, polyploidy, decreased cell density, increased cell size, increases in Descemet's thickness, and the presence of posterior projections originating from the CE towards the anterior chamber, similar to changes documented for aging human corneas. Similar changes were observed in young adult Ercc1(-/Δ) mice CE, demonstrating spontaneous premature aging of the CE of these DNA repair-deficient mice. CD45+ immune cells were associated with the posterior surface of CE from Ercc1(-/Δ) mice and the tissue expressed increased IL-1α, Cxcl2, and TNFα, pro-inflammatory proteins associated with senescence-associated secretory phenotype (SASP). These data provide strong experimental evidence that DNA damage can promote aging of the CE and that Ercc1(-/Δ) mice offer a rapid and accurate model to study CE pathogenesis and therapy. This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: To determine the retinal nerve fibre layer (RNFL) thickness at which visual field (VF) damage becomes detectable and associated with structural loss.
In a prospective cross-sectional study, 72 healthy and 40 glaucoma subjects (one eye per subject) recruited from an academic institution had VF examinations and spectral domain optical coherence tomography (SD-OCT) optic disc cube scans (Humphrey field analyser and Cirrus HD-OCT, respectively). Comparison of global mean and sectoral RNFL thicknesses with VF threshold values showed a plateau of threshold values at high RNFL thicknesses and a sharp decrease at lower RNFL thicknesses. A 'broken stick' statistical model was fitted to global and sectoral data to estimate the RNFL thickness 'tipping point' where the VF threshold values become associated with the structural measurements. The slope for the association between structure and function was computed for data above and below the tipping point.
The mean RNFL thickness threshold for VF loss was 75.3 μm (95% CI: 68.9 to 81.8), reflecting a 17.3% RNFL thickness loss from age-matched normative value. Above the tipping point, the slope for RNFL thickness and threshold value was 0.03 dB/μm (CI: -0.02 to 0.08) and below the tipping point, it was 0.28 dB/μm (CI: 0.18 to 0.38); the difference between the slopes was statistically significant (p<0.001). A similar pattern was observed for quadrant and clock-hour analysis.
Substantial structural loss (∼17%) appears to be necessary for functional loss to be detectable using the current testing methods.
The British journal of ophthalmology 04/2011; 96(1):47-52. DOI:10.1136/bjo.2010.196907 · 2.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Optical coherence tomography (OCT) imaging has become widespread in ophthalmology over the past 15 years, because of its ability to visualize ocular structures at high resolution. This article reviews the history of OCT imaging of the eye, its current status, and the laboratory work that is driving the future of the technology.
[Show abstract][Hide abstract] ABSTRACT: To investigate the longitudinal effect of optic nerve crush injury in mice by measuring retinal thickness with spectral-domain optical coherence tomography (SD-OCT).
Optic nerves of one eye from each C57Bl/6 mouse were crushed under direct visualization for 3 seconds, 1 mm posterior to the globe. The optic nerve head (ONH) was imaged with SD-OCT (1.5 × 1.5 × 2.0 mm scan) before the surgical intervention and repeated subsequently for up to 32 days postinjury. A cohort of mice not exposed to the nerve crush procedure served as control. En face SD-OCT images were used to manually align subsequent scans to the baseline en face image. Total retinal thickness (TRT) (along a sampling band with radii 0.33-0.42 mm centered on the ONH) from each follow-up day was automatically quantified for global and sectoral measurements using custom software. Linear mixed-effects models with quadratic terms were fitted to compare TRT of nerve-crushed and control eyes over time.
Eleven eyes from 11 nerve crush mice (baseline age 76 ± 11.8 days) and eight eyes from four healthy mice (baseline age 64 ± 0 days) were included. The control eyes showed a small, gradual, and consistent TRT increase throughout follow-up. Nerve-crushed eyes showed an initial period of thickening, followed by thinning and slight rebound after day 21. The decrease in thickness observed after the early thickening resolved was statistically significantly different from the control eyes (P < 0.05 for global and sectoral measurements).
SD-OCT can be used to quantitatively monitor changes in retinal thickness in mice over time.
[Show abstract][Hide abstract] ABSTRACT: To test the reproducibility of spectral-domain optical coherence tomography (SD-OCT) total retinal thickness (TRT) measurements in mice.
C57Bl/6 mice were anesthetized, and three repeated volumetric images were acquired in both eyes with SD-OCT (250 A-scans × 250 frames × 1024 samplings), centered on the optic nerve head (ONH). The mice were repositioned between scans. TRT was automatically measured within a sampling band of retinal thickness with radii of 55 to 70 pixels, centered on the ONH by using custom segmentation software. The first volumetric image acquired in a given eye was used to register the remaining two SD-OCT images by manually aligning the en face images with respect to rotation and linear translation. Linear mixed-effects models were fitted to global and quadrant thicknesses, taking into account the clustering between eyes, to assess imprecision (measurement reproducibility).
Twenty-six eyes of 13 adult mice (age 13 weeks) were imaged. The mean global TRT across all eyes was 298.21 μm, with a mouse heterogeneity standard deviation (SD) of 4.88 μm (coefficient of variation [CV] = 0.016), an eye SD of 3.32 μm (CV = 0.011), and a device-related imprecision SD of 2.33 μm (CV = 0.008). The superior quadrant had the thickest mean TRT measurement (310.38 μm) and the highest (worst) imprecision SD (3.13 μm; CV = 0.010), and the inferior quadrant had the thinnest mean TRT (291.55 μm). The quadrant with the lowest (best) imprecision SD was in the nasal one (2.06 μm; CV = 0.007).
Good reproducibility was observed for SD-OCT retinal thickness measurements in mice. SD-OCT may be useful for in vivo longitudinal studies in mice.
[Show abstract][Hide abstract] ABSTRACT: Three dimensional (3D) ophthalmic imaging using optical coherence tomography (OCT) has revolutionized assessment of the eye, the retina in particular. Recent technological improvements have made the acquisition of 3D-OCT datasets feasible. However, while volumetric data can improve disease diagnosis and follow-up, novel image analysis techniques are now necessary in order to process the dense 3D-OCT dataset. Fundamental software improvements include methods for correcting subject eye motion, segmenting structures or volumes of interest, extracting relevant data post hoc and signal averaging to improve delineation of retinal layers. In addition, innovative methods for image display, such as C-mode sectioning, provide a unique viewing perspective and may improve interpretation of OCT images of pathologic structures. While all of these methods are being developed, most remain in an immature state. This review describes the current status of 3D-OCT scanning and interpretation, and discusses the need for standardization of clinical protocols as well as the potential benefits of 3D-OCT scanning that could come when software methods for fully exploiting these rich datasets are available clinically. The implications of new image analysis approaches include improved reproducibility of measurements garnered from 3D-OCT, which may then help improve disease discrimination and progression detection. In addition, 3D-OCT offers the potential for preoperative surgical planning and intraoperative surgical guidance.
Progress in Retinal and Eye Research 11/2010; 29(6):556-79. DOI:10.1016/j.preteyeres.2010.05.005 · 8.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Measurements of human Schlemm's canal (SC) have been limited to histologic sections. The purpose of this study was to demonstrate noninvasive measurements of aqueous outflow (AO) structures in the human eye, examining regional variation in cross-sectional SC areas (on/off collector channel [CC] ostia [SC/CC] and nasal/temporal) in the eyes of living humans.
SC was imaged by spectral-domain optical coherence tomography with a 200-nm bandwidth light source. Both eyes of 21 healthy subjects and one glaucomatous eye of three subjects were imaged nasally and temporally. Contrast and magnification were adjusted to maximize visualization. Cross-sectional SC on and off SC/CC was traced three times by two independent masked observers using ImageJ (ImageJ 1.40g, http://rsb.info.nih.gov/ij/ Wayne Rasband, developer, National Institutes of Health, Bethesda, MD). The mean SC area was recorded. A linear mixed-effects model was used to analyze eye, nasal/temporal laterality, and SC area on or off SC/CC.
SC area was significantly larger on SC/CCs than off (12,890 vs. 7,391 micorm(2), P < 0.0001) and was significantly larger on the nasal side than on the temporal (10,983 vs. 8,308 micorm(2), P = 0.009). SC areas were significantly smaller in glaucoma patients than in normal subjects, whether pooled (P = 0.0073) or grouped by on (P = 0.0215) or off (P = 0.0114) SC/CC.
Aqueous outflow structures, including SC and CCs, can be noninvasively assessed in the human eye. These measurements will be useful in physiological studies of AO and will be clinically useful in the determination of the impact of glaucoma therapies on IOP as well as presurgical planning.
[Show abstract][Hide abstract] ABSTRACT: Optical coherence tomography (OCT) provides real-time, objective,
in-vivo, optical cross-sectional representations of the retina and optic
nerve. Recent innovations in image acquisition, including the
incorporation of Fourier/spectral-domain detection, have improved
imaging speed, sensitivity and resolution. Still, there remain specific
structures within ocular OCT images, such as retinal ganglion cells
(RGCs), which are of clinical interest but consistently have low
contrast. This makes it difficult to differentiate between surrounding
layers and structures. The objectives of this project were: (1) To
establish a reliable method for OCT imaging of the healthy and diseased
mouse eye in order to provide a platform for testing the utility of OCT
contrast agents for ocular imaging, (2) To develop antibody-conjugated
gold nanoparticles suitable for targeting specific structures and
enhancing OCT image contrast in the mouse eye, and (3) To examine the
localized contrast-enhancing ability and biocompatibility of gold
nanoparticle contrast agents in-vivo. Our organizing hypotheses were
that nanoparticles could improve contrast by modulating the intensity of
backscattered light detected by OCT and that they could be directed to
ocular structures of interest using antibodies specific to cellular
markers. A reproducible method for imaging the mouse retina and
quantifying retinal thickness was developed and this technique was then
applied to a mouse model for retinal ganglion cell loss, optic nerve
crush. Gold nanorods were designed specifically to augment the
backscattering OCT signal at the same wavelengths of light used in
current ophthalmic OCT imaging schemes (resonant wavelength lambda = 840
nm). Anti-CD90.1 (Thy1.1) antibodies were conjugated to the gold
nanorods and a protocol for characterization of the success of antibody
conjugation was developed. Upon injection, the gold nanorods were found
to remain in the vitreous post-injection, with many consumed by an early
inflammatory response and only very few reaching the internal limiting
membrane and passing into the retina. Our findings suggest that, while
gold nanorods are able to locally increase OCT signal intensity in the
vitreous, their utility in the retina may be limited.
[Show abstract][Hide abstract] ABSTRACT: Time domain optical coherence tomography (TD-OCT) has been used commonly in clinical practice, producing a large inventory of circular scan data for retinal nerve fiber layer (RNFL) assessment. Spectral domain (SD)-OCT produces three-dimensional (3-D) data volumes. The purpose of this study was to create a robust technique that makes TD-OCT circular scan RNFL thickness measurements comparable with those from 3-D SD-OCT volumes.
Eleven eyes of 11 healthy subjects and 7 eyes of 7 subjects with glaucoma were enrolled. Each eye was scanned with one centered and eight displaced TD-OCT scanning circles. One 3-D SD-OCT cube scan was obtained at the same visit. The matching location of the TD-OCT scanning circle was automatically detected within the corresponding 3-D SD-OCT scan. Algorithm performance was assessed by estimating the difference between the detected scanning circle location on 3-D SD-OCT volume and the TD-OCT circle location. Global and sectoral RNFL thickness measurement errors between the two devices were also compared.
The difference (95% confidence interval) in scanning circle center locations between TD- and SD-OCT was 2.3 (1.5-3.2) pixels (69.0 [45.0-96.0] microm on the retina) for healthy eyes and 3.1 (2.0-4.1) pixels (93.0 [60.0-123.0] microm on the retina) for glaucomatous eyes. The absolute RNFL thickness measurement difference was significantly smaller with the matched scanning circle.
Scan location matching may bridge the gap in RNFL thickness measurements between TD-OCT circular scan data and 3-D SD-OCT scan data, providing follow-up comparability across the two generations of OCTs.
[Show abstract][Hide abstract] ABSTRACT: To evaluate, within ocular imaging scans of acceptable quality as determined by manufacturers' guidelines, the effects of image quality on glaucoma discrimination capabilities.
One hundred and four healthy and 75 glaucomatous eyes from the Advanced Imaging in Glaucoma Study (AIGS) were imaged with GDx-VCC, HRT II and StratusOCT. Quality score (QS>/=8), pixel standard deviation (SD</=50) and signal strength (SS>/=5) were used as quality parameter cut-offs, respectively. GDx nerve fibre indicator (NFI) and HRT Moorfields regression analysis (MRA) classifications and OCT mean retinal nerve fibre layer (RNFL) thickness were used as the discriminatory parameters. Logistic regression models were used to model the dichotomous clinical classification (healthy vs glaucoma) as a function of image-quality parameters and discriminatory parameters.
Quality parameter covariates were statistically non-significant for GDx and HRT but had an inverse effect on OCT in predicting disease (a higher SS had a lower probability of glaucoma). Age was a significant covariate for GDx and HRT, but not OCT, while ethnicity and interaction between the image quality and the institute where scans were acquired were significant covariates in the OCT models.
Scan quality within the range recommended as acceptable by the manufacturer of each imaging device does not affect the glaucoma discriminating ability of GDx or HRT but does affect Stratus OCT glaucoma discrimination.
The British journal of ophthalmology 08/2009; 93(12):1580-4. DOI:10.1136/bjo.2008.152223 · 2.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate retinal nerve fibre layer (RNFL) thickness measurement reproducibility using conventional time-domain optical coherence tomography (TD-OCT) and spectral-domain OCT (SD-OCT), and to evaluate two methods defining the optic nerve head (ONH) centring: Centred Each Time (CET) vs Centred Once (CO), in terms of RNFL thickness measurement variability on SD-OCT.
Twenty-seven eyes (14 healthy subjects) had three circumpapillary scans with TD-OCT and three raster scans (three-dimensional or 3D image data) around ONH with SD-OCT. SD-OCT images were analysed in two ways: (1) CET: ONH centre was defined on each image separately and (2) CO: ONH centre was defined on one image and exported to other images after scan registration. After defining the ONH centre, a 3.4 mm diameter virtual circular OCT was resampled on SD-OCT images to mimic the conventional circumpapillary RNFL thickness measurements taken with TD-OCT.
CET and CO showed statistically significantly better reproducibility than TD-OCT except for 11:00 with CET. CET and CO methods showed similar reproducibility.
SD-OCT 3D cube data generally showed better RNFL measurement reproducibility than TD-OCT. The choice of ONH centring methods did not affect RNFL measurement reproducibility.
The British journal of ophthalmology 06/2009; 93(8):1057-63. DOI:10.1136/bjo.2009.157875 · 2.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Optical coherence tomography has allowed unprecedented visualization of ocular structures, but the identity of some visible objects within slices remains unknown. This study reconstructs a number of those objects in 3D space, allowing their identification by observation of their 3D morphology. In the case mottling deep within image slices through the optic disc, C-mode imaging provided visualization of the appearance and distribution of laminar pores. In the case of white spots and streaks sometimes observed in image slices through the cornea, C-mode imaging contoured to the path of those white spots allowed their visual identification as nerves extending radially into the cornea from the limbus. White spots observed in ultra-high resolution retinal image slices were identified as blood within retinal capillaries. C-mode contour-corrected imaging of three dimensional structures provided the identification of previously unidentified structures visible in cross-sectional image slices.
[Show abstract][Hide abstract] ABSTRACT: To develop a semiautomated method to visualize structures of interest (SoIs) along their contour within three-dimensional, spectral domain optical coherence tomography (3D SD-OCT) data, without the need for segmentation.
With the use of two SD-OCT devices, the authors obtained 3D SD-OCT data within 6 x 6 x 1.4-mm and 6 x 6 x 2-mm volumes, respectively, centered on the fovea in healthy eyes and in eyes with retinal pathology. C-mode images were generated by sampling a variable thickness plane semiautomatically modeled to fit the contour of the SoI. Unlike published and commercialized methods, this method did not require retinal layer segmentation, which is known to fail frequently in the presence of retinal pathology. Four SoIs were visualized for healthy eyes: striation of retinal nerve fiber (RNF), retinal capillary network (RCN), choroidal capillary network (CCN), and major choroidal vasculature (CV). Various SoIs were visualized for eyes with retinal pathology.
Seven healthy eyes and seven eyes with retinal pathology (cystoid macular edema, central serous retinopathy, vitreoretinal traction, and age-related macular degeneration) were imaged. CCN and CV were successfully visualized in all eyes, whereas RNF and RCN were visualized in all healthy eyes and in 42.8% of eyes with pathologies. Various SoIs were successfully visualized in all eyes with retinal pathology.
The proposed C-mode contour modeling may provide clinically useful images of SoIs even in eyes with severe pathologic changes in which segmentation algorithms fail.
[Show abstract][Hide abstract] ABSTRACT: Study of the structure of the lamina cribrosa is critical in glaucoma research. The purpose of this study is to determine the optimal spectral domain optical coherence tomography imaging protocol for the digital isolation and display of the lamina cribrosa. Three-dimensional datasets centered on the lamina cribrosa were obtained with 200 X 200 to 512 X 512 A-scan densities. The effect of scan density and c-mode slab thickness was subjectively compared. Increasing slab thickness reduced the sharpness of visible prelamina and lamina cribrosa structures. In retrolamina structures, thin slabs provided good visualization, but increased slab size increased the visibility of deeper structures. Scan times as short as 2.3 seconds (256 X 256 A-scans) degraded visualization of the shape of the optic nerve head. The optical scan protocol for lamina cribrosa imaging appears to be a 3 x 3 mm 200 X 200 A-scan volume with the lamina cribrosa positioned near direct current.
Ophthalmic Surgery Lasers and Imaging 07/2008; 39(4 Suppl):S126-131. · 1.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the effect on optical coherence tomography (OCT) retinal nerve fiber layer (RNFL) thickness measurements of varying the standard 3.4-mm-diameter circle location.
The optic nerve head (ONH) region of 17 eyes of 17 healthy subjects was imaged with high-speed, ultrahigh-resolution OCT (hsUHR-OCT; 501 x 180 axial scans covering a 6 x 6-mm area; scan time, 3.84 seconds) for a comprehensive sampling. This method allows for systematic simulation of the variable circle placement effect. RNFL thickness was measured on this three-dimensional dataset by using a custom-designed software program. RNFL thickness was resampled along a 3.4-mm-diameter circle centered on the ONH, then along 3.4-mm circles shifted horizontally (x-shift), vertically (y-shift) and diagonally up to +/-500 microm (at 100-microm intervals). Linear mixed-effects models were used to determine RNFL thickness as a function of the scan circle shift. A model for the distance between the two thickest measurements along the RNFL thickness circular profile (peak distance) was also calculated.
RNFL thickness tended to decrease with both positive and negative x- and y-shifts. The range of shifts that caused a decrease greater than the variability inherent to the commercial device was greater in both nasal and temporal quadrants than in the superior and inferior ones. The model for peak distance demonstrated that as the scan moves nasally, the RNFL peak distance increases, and as the circle moves temporally, the distance decreases. Vertical shifts had a minimal effect on peak distance.
The location of the OCT scan circle affects RNFL thickness measurements. Accurate registration of OCT scans is essential for measurement reproducibility and longitudinal examination (ClinicalTrials.gov number, NCT00286637).
[Show abstract][Hide abstract] ABSTRACT: The purpose of this study was to compare the day-to-day reproducibility of optical coherence tomography (OCT; StratusOCT, Carl Zeiss Meditec, Dublin, CA) measurements of retinal nerve-fibre layer (RNFL) measurements at time points 1 year apart.
One eye in each of 11 healthy subjects was examined using the StratusOCT fast RNFL scan protocol. Three fast RNFL scans with signal strength > or =7 were obtained on each of 3 days within a month. This protocol was repeated after 12 months. A linear mixed effects model fitted to the nested data was used to compute the variance components.
The square root of the variance component that was attributed to the differences between subjects was 7.17 microm in 2005 and 7.28 microm in 2006. The square roots of the variance component due to differences between days within a single subject were 1.95 microm and 1.50 microm, respectively, and for within day within a single subject were 2.51 microm and 2.55 microm, respectively. There were no statistically significant differences for any variance component between the two testing occasions.
Measurement error variance remains similar from year to year. Day and scan variance component values obtained in a cohort study may be safely applied for prediction of long-term reproducibility.
The British journal of ophthalmology 06/2008; 92(6):806-9. DOI:10.1136/bjo.2007.129312 · 2.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To assess performance of classifiers trained on Heidelberg Retina Tomograph 3 (HRT3) parameters for discriminating between healthy and glaucomatous eyes.
Classifiers were trained using HRT3 parameters from 60 healthy subjects and 140 glaucomatous subjects. The classifiers were trained on all 95 variables and smaller sets created with backward elimination. Seven types of classifiers, including Support Vector Machines with radial basis (SVM-radial), and Recursive Partitioning and Regression Trees (RPART), were trained on the parameters. The area under the ROC curve (AUC) was calculated for classifiers, individual parameters and HRT3 glaucoma probability scores (GPS). Classifier AUCs and leave-one-out accuracy were compared with the highest individual parameter and GPS AUCs and accuracies.
The highest AUC and accuracy for an individual parameter were 0.848 and 0.79, for vertical cup/disc ratio (vC/D). For GPS, global GPS performed best with AUC 0.829 and accuracy 0.78. SVM-radial with all parameters showed significant improvement over global GPS and vC/D with AUC 0.916 and accuracy 0.85. RPART with all parameters provided significant improvement over global GPS with AUC 0.899 and significant improvement over global GPS and vC/D with accuracy 0.875.
Machine learning classifiers of HRT3 data provide significant enhancement over current methods for detection of glaucoma.
The British journal of ophthalmology 06/2008; 92(6):814-8. DOI:10.1136/bjo.2007.133074 · 2.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To test if improving optical coherence tomography (OCT) resolution and scanning speed improves the visualization of glaucomatous structural changes as compared with conventional OCT.
Prospective observational case series.
Healthy and glaucomatous subjects in various stages of disease.
Subjects were scanned at a single visit with commercially available OCT (StratusOCT) and high-speed ultrahigh-resolution (hsUHR) OCT. The prototype hsUHR OCT had an axial resolution of 3.4 mum (3 times higher than StratusOCT), with an A-scan rate of 24 000 hertz (60 times faster than StratusOCT). The fast scanning rate allowed the acquisition of novel scanning patterns such as raster scanning, which provided dense coverage of the retina and optic nerve head.
Discrimination of retinal tissue layers and detailed visualization of retinal structures.
High-speed UHR OCT provided a marked improvement in tissue visualization as compared with StratusOCT. This allowed the identification of numerous retinal layers, including the ganglion cell layer, which is specifically prone to glaucomatous damage. Fast scanning and the enhanced A-scan registration properties of hsUHR OCT provided maps of the macula and optic nerve head with unprecedented detail, including en face OCT fundus images and retinal nerve fiber layer thickness maps.
High-speed UHR OCT improves visualization of the tissues relevant to the detection and management of glaucoma.
[Show abstract][Hide abstract] ABSTRACT: To compare stereometric parameters and classification results from the Heidelberg Retina Tomograph version 2 (HRT2); HRT3; and HRT3 Glaucoma Probability Score (GPS), an automated method of obtaining optic nerve head analysis without the need for manual definition of disc margin.
Retrospective cross-sectional study.
Five hundred four eyes from 281 consecutive subjects (glaucoma, glaucoma suspect, and healthy) evaluated in a glaucoma clinic.
All participants had HRT2 scanning of the optic nerve head. Inclusion criteria were scans with good centration and focus, even illumination, an overall quality score by HRT3 of acceptable or better, and standard deviation < 50 mum. A Bland-Altman analysis was used for the comparison of HRT2 and HRT3. From these results, calibration equations were determined to permit conversion of the measurements between devices. The agreement between HRT2 and HRT3 Moorfields regression analysis (MRA) and HRT3 GPS classification methods was measured using kappa statistics.
Heidelberg Retina Tomograph version 2 and HRT3 stereometric parameters, MRA, and global GPS.
There was a statistically significant difference between HRT2 and HRT3 global disc area, rim area, cup area, rim volume, cup volume, height variation contour, and retinal nerve fiber layer cross-sectional area stereometric parameters. All of those parameters were smaller using HRT3, due to a manufacturer-reported horizontal scaling error of 4% in HRT2 that was corrected in HRT3. kappas for agreement were 0.60 between classifications (within normal limits, borderline, and outside normal limits) of MRA by HRT2 and HRT3 and 0.47 between HRT3 MRA and GPS.
The HRT3 generally provided smaller stereometric disc measurements than HRT2. There was no clear conversion between HRT3 and GPS parameters, as the 2 methods for measuring the stereometric parameters differ.
[Show abstract][Hide abstract] ABSTRACT: To demonstrate a new imaging method for high resolution spectral domain optical coherence tomography (SD-OCT) for small animal developmental imaging.
Wildtype zebrafish that were 24, 48, 72, and 120 h post fertilization (hpf) and nok gene mutant (48 hpf) embryos were imaged in vivo. Three additional embryos were imaged twice, once at 72 hpf and again at 120 hpf. Images of the developing eye, brain, heart, whole body, proximal yolk sac, distal yolk sac, and tail were acquired. Three-dimensional OCT data sets (501 x 180 axial scans) were obtained as well as oversampled frames (8,100 axial scans) and repeated line scans (180 repeated frames). Scan volumes ranged from 750 x 750 microm to 3 x 3 mm, each 1.8 mm thick. Three-dimensional data sets allowed construction of C-mode slabs of the embryo.
SD-OCT provided ultra-high resolution visualization of the eye, brain, heart, ear, and spine of the developing embryo as early as 24 hpf, and allowed development to be documented in each of these organ systems in consecutive sessions. Repeated line scanning with averaging optimized the visualization of static and dynamic structures contained in SD-OCT images. Structural defects caused by a mutation in the nok gene were readily observed as impeded ocular development, and enlarged pericardial cavities.
SD-OCT allowed noninvasive, in vivo, ultra-high resolution, high-speed imaging of zebrafish embryos in their native state. The ability to measure structural and functional features repeatedly on the same specimen, without the need to sacrifice, promises to be a powerful tool in small animal developmental imaging.