In this presentation we will evaluate the signal processing steps required to obtain stimulus-evoked nm-scale changes in the photoreceptor outer segment (OS) length using full field swept source optical coherence tomography (FF-SS-OCT). Evaluation of stimulus-evoked OS length changes is the basis of photoreceptor optoretinography (ORG), an emerging test of retinal function. To allow cellular resolution, an adaptive optics (AO) enhanced FF-SS-OCT system was constructed. This system allows up to kHz volume acquisition rates, which greatly reduce retinal motion artifacts. Specifically, the effects of wave number calibration, dispersion compensation, and application of short-time Fourier transform (STFT) on image quality metrics, SNR of individual OCT volumes, and extracted phase noise will be presented.
While the a-wave of mouse electroretinogram (ERG) occurs within 50 milliseconds after exposure to light, the optoretinogram (ORG) slower than a 20Hz sampling rate could face limitations in observing immediate morphological changes from the retina. In this study, we develop a compact custom-built mouse ORG system based on spectral domain optical coherence tomography (SD-OCT) for 100Hz~1KHz B-scan rates comprised of 100 kHz A-scans. All the optics of the developed ORG system are designed on a 24 x 24 inches optical breadboard to move easily as well as to combine with the ERG system in a dark room. Without using a fundus camera, the OCT system provides en-face images from high-pass filtering and square of the OCT spectral signal for mouse retinal positioning in-vivo before acquiring ORG data. The 490nm LED for light stimulus is generated to make uniform illumination at the mouse retina using the Maxwellian view method. The common path of the OCT scanning light and the visible LED is built with achromatic doublet lens combinations based on optical simulation with Opticstudio® . The developed compact ORG system can not only observe light-evoked responses with 1~10 milliseconds but also be used for the studies of correlations between ORG and ERG in the mouse retina.
Observing micro-vessels in conjunctiva could be used not only for diagnosing conjunctival diseases including conjunctivitis and pterygium but also as biomarkers for circulatory diseases. Many research teams have developed compact imaging and auto-analysis systems to simplify the conventional slit lamp as well as enhance the analysis process. The imaging system, previously developed by our research team, corrects eye motion in imaging windows through image registration and template matching. The developed system quantifies blood flow velocity using the sequence of motion-corrected images. This study compares estimated flow velocity and the fluid's actual velocity using the experimental phantom comprising transparent hoses and fluid including beads corresponding to red blood cells. The flow velocities are calculated using the Hagen-Poiseuille equation, and the flow rates generated by the syringe pump. The pump applies three kinds of flow rates to generate flow velocity variations and the estimated velocities are linearly proportional to these variations. In addition, the phantom has a random motion to mimic the fixational eye movements within the range of the healthy subject's angular eye motion. Through these experiments, we verified the previously developed flow velocity measurement system having percent errors under 3% by comparing estimated flow velocities with actual flow velocities. The system's accuracy, especially under conditions without artificial motion, is over 98.5%. These experiments can provide the supporting background for feasibility and accuracy in a further clinical study in conjunctival microcirculation.
Adaptive optics(AO) compensates for aberration between light detector and imaging target, for example, air turbulence, misaligned optics, and eye lens. This technique, initially developed to improve the performance of astronomic imaging, is leading to advances in the ophthalmic imaging field through combination with various optical imaging systems. General hardware-based AO systems need additional optics and beam paths for adjusting the beam size at the deformable mirror(DM) as well as matching the image plane. Previous research has composed the system using spherical mirrors instead of the lens to reduce aberration. Additionally, previous papers have reported off-axis type AO imaging systems for eliminating astigmatism and more compact equipment than on-axis type. This study optimized an off-axis AO-region of the AO-scanning laser ophthalmoscope(SLO) through optical design by Opticstudio® and three-dimensional rendering by SOLIDWORKS®. The rendering process, including virtual mounting, allows checking whether block or pass of the beam by optomechanics and providing relative coordinate of mounted optics. The verification method for checking the alignment of the system is the comparison between simulation and practical beam wavefront at a specific beam path.
Microgravity, vacuum, and high-intensity ultraviolet waves are widely known characteristics of space. These different environments from the earth affect physical changes including ocular tissue changes while astronauts stay in the universe. The changes in ocular tissue in the space environment, also known as visual impairment intracranial pressure (VIIP) syndrome, including fundus optic disc edema, hyperopic drift, choroidal folds, cotton spots, and permanent fundus damage could influence astronauts’ vision system and ability of space operations. Especially, hyperopic drift by posterior flattening and folded retina by choroidal folds are reported to affect the retina's structures as a vision sensor directly. To investigate microgravity's effect on ocular tissues and vision, previous research on earth are used special facilities and various microgravity simulators, including head-down tilt bed-rest and random positioning machines. This study suggests that an experiment expose wild-type zebrafish to microgravity using a rotary cell culture system (RCCs) applied to experiments using cell and zebrafish's embryos in microgravity. Unlike previous research using zebrafish's embryos and larva, adult and growing zebrafish were employed in this study for observing ocular changes in simulated space environments. After exposing zebrafish to microgravity, in-vivo zebrafish's eye images were acquired by custom-built optical coherence tomography (OCT). This research for presenting the new method for small animal experiments in microgravity environments could be applied to investigate the influence of staying in the universe on an animal model with ophthalmic diseases.
KEYWORDS: Retina, 3D image processing, Stereoscopy, In vivo imaging, Optical coherence tomography, Injuries, Eye models, Eye, Animal model studies, Analytical research
The retina regeneration process has been observed in several animals including fish, birds, and amphibians, whereas the injured human retina cannot regenerate until today. The zebrafish is one of the animals which can regenerate their retina. Due to similarities between humans and the zebrafish in the morphology of the eye as well as the gene, the zebrafish has been chosen as a useful model for investigating retina regeneration. Previous studies have observed morphological changes in the zebrafish retina using optical coherence tomography during the retinal regeneration after light irradiation with a single beam power. However, these studies have limitations of demonstrating differences in regenerative abilities depending on degrees of retinal damages because they used only a single light irradiating condition. Through light-induced retinal injuries by various light irradiating conditions, investigation correlations between the time required for retina regeneration and grades of retinal damages were needed. To conduct these experiments, a custom-built OCT, which can acquire a cross-sectional and three-dimensional image of the zebrafish's eye, was developed based on OpticStudio simulations. The zebrafish's eye imaging can provide the ability to observe the damage and morphological changes after laser irradiation in the eye tissues including the cornea and retina.
This paper presents the preliminary study on monitoring of intraocular pressure (IOP) of human eye by using optical coherence tomography (OCT). Because hypertension IOP can lead to Glaucoma, one of chronic diseases of the optic nerve, the periodic monitoring of IOP through tonometry is essential to prevent the Glaucoma. One of the most common tonometry methods to estimate IOP includes measuring corneal deflection by using either a direct contact or non-contact (e.g. air puff) impact force. However, this approach may be harmful to human eye. Therefore, it is necessary to develop the more reliable and safe monitoring method. In ophthalmology, the differentiation of open-angle glaucoma and normal eyes is used to monitor the IOP by using. Typically, peripapillary vessel density or superficial perifoveal macular vessel density measurements are currently utilized. In this study, optical OCT images of bovine eye was captured with different IOPs of bovine and analyzed to extract the signature where defines the relationship between IOP and optical OCT images.
Low-level light therapy, which is a red or near-infrared light-employed therapeutic methodology, can act to help the brain repair in cases of traumatic brain injury and stroke. In this paper, we investigate the effects of near-infrared light therapy (NILT) for the recovery of blood flow of mice with cerebral hypoperfusion, which is a key mechanism leading to vascular dementia, induced by bilateral common carotid artery stenosis (BCAS). The mice are divided into three groups of 4-5 mice per group: a normal group (without BCAS operation), a BCAS group (without NILT), or a BCAS/NILT group. The LED with a peak wavelength of 810 nm and a power of 20 mW is arranged to illuminate on the top of the mouse head. The mice receive treatment from the LED source 3 times per week for one month. After the treatment, positron emission tomography is used to quantify the effects of NILT on whole brain and regional cerebral blood flow in the cortex, striatum, and hippocampus.
Dermatologic patients have various skin characteristics such as skin tone and pigmentation color. However most studies on laser ablation and treatment only considered laser operating conditions like wavelength, output power and pulse duration. The laser ablation arises from photothermal effect by photon energy absorption. Chromophores like melanin exist as the absorber in the skin. In this study, we painted color to mimic chromophores on in-vivo and in-vitro skin models to demonstrate influence on the laser ablation by skin color. Water-based pens were used to paint color. Cross sectional images of the laser ablation were acquired by Fourier-domain optical coherence tomography (Fd-OCT). Light source to make ablation was a Q-switch diode-pumped Nd:YVO4 nanosecond laser (532nm central wavelength). Irradiated light energy dose of the laser could not make ablation craters in the control group. However experimental groups showed craters with same irradiation light energy dose. These results show painting on skin increased tissue damage by absorption in painted color without dyeing cells or tissues.
Laser-induced therapies include laser ablation to remove or cut target tissue by irradiating high-power focused laser beam. These laser treatments are widely used tools for minimally invasive surgery and retinal surgical procedures in clinical settings. In this study, we demonstrate laser tissue interaction images of various sample tissues using high resolution Fourier-domain optical coherence tomography (Fd-OCT). We use a Q-switch diode-pumped Nd:YVO4 nanosecond laser (532nm central wavelength) with a 4W maximum output power at a 20 kHz repetition rate to ablate in vitro and in vivo samples including chicken breast and mouse ear tissues. The Fd-OCT system acquires time-series Bscan images at the same location during the tissue ablation experiments with 532nm laser irradiation. The real-time series of OCT cross-sectional (B-scan) images compare structural changes of 532nm laser ablation using same and different laser output powers. Laser tissue ablation is demonstrated by the width and the depth of the tissue ablation from the B-scan images.
We report on development of optical parametric oscillator (OPO) based mid-infrared laser system, which utilizes periodically poled nonlinear crystal that was pumped by near-infrared (NIR) laser. We have obtained 8 W of mid-infrared average output at the injection current of 20A from a quasi-phase-matched OPO using external cavity configuration. The laser tissue ablation efficiency was investigated which is substantially affected by several parameters such as, optical fluence rate, wavelength of the laser source and the optical properties of target tissue. Wavelength and radiant exposure dependent tissue ablation dimension were quantified by using SD-OCT (spectral domain optical coherence tomography) and the ablation efficiency was compared to that of non-converted NIR- laser system.
Low-level light irradiation (LLLI) reported to stimulate the proliferation or differentiation of a variety of cell types. However, very little is known about the effect of light therapy on stem cells. The aim of the present study was to evaluate the effect of LLLI on the molecular physiological change of human bone marrow derived stem cells (hBMSC) by wavelength (470, 630, 660, 740 and 850, 50mW). The laser diode was performed with different time interval (0, 7.5, 15, 30J/cm2, 50mW) on hBMSC. To determine the molecular physiological changes of cellular level of hBMSC, the clonogenic assay, ATP assay, reactive oxygen species (ROS) detection, mitochondria membrane potential (MMPΦ) staining and calcium efflux assay were assessed after irradiation. There was a difference between with and without irradiation on hBMSCs. An energy density up to 30 J/cm² improved the cell proliferation in comparison to the control group. Among these irradiated group, 630 and 660nm were significantly increased the cell proliferation. The cellular level of ATP and calcium influx was increased with energy dose-dependent in all LLLI groups. Meanwhile, ROS and MMPΦ were also increased after irradiation except 470nm. It can be concluded that LLLI using infrared light and an energy density up to 30 J/cm² has a positive stimulatory effect on the proliferation or differentiation of hBMSCs. Our results suggest that LLLI may influence to the mitochondrial membrane potential activity through ATP synthesis and increased cell metabolism which leads to cell proliferation and differentiation.
We present a noninvasive phase-variance (pv)–based motion contrast method for depth-resolved imaging of the human chorioretinal complex microcirculation with a newly developed phase-stabilized high speed (100-kHz A-scans/s) 1-μm swept-source optical coherence tomography (SSOCT) system. Compared to our previous spectral-domain (spectrometer based) pv-spectral domain OCT (SDOCT) system, this system has the advantages of higher sensitivity, reduced fringe wash-out for high blood flow speeds and deeper penetration in choroid. High phase stability SSOCT imaging was achieved by using a computationally efficient phase stabilization approach. This process does not require additional calibration hardware and complex numerical procedures. Our phase stabilization method is simple and can be employed in a variety of SSOCT systems. Examples of vasculature in the chorioretinal complex imaged by pv-SSOCT from normal as well as diseased eyes are presented and compared to retinal images of the same subjects acquired with fluorescein angiography and indocyanine green angiography. Observations of morphology of vascular perfusion in chorioretinal complex visualized by our method are listed.
We demonstrate the feasibility of our newly developed phase stabilized high-speed (100 kHz A-scans/s) 1 μm sweptsource
optical coherence tomography (SSOCT) system with the phase-variance based motion contrast method for
visualization of human chorioretinal complex microcirculation. Compared to our previously reported spectral domain
(spectrometer based) phase-variance (pv)-SDOCT system it has advantages of higher sensitivity, reduced fringe washout
for high blood flow speeds and deeper penetration in choroid. High phase stability SSOCT imaging was achieved by
using a computationally efficient phase stabilization approach. This process does not require additional calibration
hardware and complex numerical procedures. Our phase stabilization method is simple and can be employed in a variety
of SS-OCT systems. Examples of vasculature in the chorioretinal complex imaged by pv-SSOCT is presented and
compared to retinal images of the same volunteers acquired with fluorescein angiography (FA) and indocyanine green
angiography (ICGA).
We present in vivo noninvasive retinal and choroidal perfusion maps with phase-variance optical coherence tomography (pvOCT). We acquired a pvOCT volumetric data set of a normal subject and visualized blood circulation in the retina and the choroid. En face projection views of the retina as well as the choroid were generated from a manually segmented volumetric data set. In addition, the processed pvOCT images were compared to current standard imaging modalities used for retinal and choroidal vasculature visualization in clinical settings, including fluorescein angiography (FA) and indocyanine green angiography (ICGA).
We acquired in vivo images of the human optic nerve head (ONH) using an adaptive optics—optical coherence tomography (AO-OCT) system. In order to improve imaging of the lamina cribrosa in the ONH with high lateral resolution and sensitivity, we implemented a complex conjugate artifact-free Fourier domain OCT (Fd-OCT) acquisition scheme with a reference arm-based phase shifting method. This allowed positioning of the lamina cribrosa structures near the zero path length difference where AO-OCT imaging achieves highest sensitivity. Implementation of our complex conjugate artifact removal (CCR) method required constant phase shifts between consecutive axial scans (A-scans), generated by continuous beam path-length changes from offsetting the pivot point of the scanning mirror placed in the reference arm. Fourier transform along the transverse axis and a filtering algorithm allowed reconstruction of CCR AO-OCT images. The suppression ratio of the mirror artifact was approximately 22 dB (at 18,000 A-scans per second acquisition speed) with a paperboard test target and an optimum phase-shift value. Finally, we reconstructed the three-dimensional structure of human ONH with enhanced depth range and sensitivity using CCR AO-OCT.
Recent progress in retinal image acquisition techniques, including optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO), combined with improved performance of adaptive optics (AO) instrumentation, has resulted in improvement in the quality of in vivo images of cellular structures in the outer layers of the human retina. Despite the significant progress in imaging cone and rod photoreceptor mosaics, visualization of cellular structures in the
inner retina has been achieved only with extrinsic contrast agents that have not been approved for use with humans. In this paper we describe the main limiting factors in visualizing inner retinal cells and the methods we implemented to reduce their effects on images acquired with AO-OCT. These include improving the system point spread function (AO performance), monitoring of motion artifacts (retinal motion tracking), and speckle pattern reduction (temporal and spatial averaging). Results of imaging inner retinal morphology and the improvement offered by the new UC Davis AOOCT
system with spatio-temporal image averaging are presented.
KEYWORDS: Optical coherence tomography, Retina, Image segmentation, Visualization, Capillaries, Image processing, Doppler tomography, Data acquisition, In vivo imaging, Data processing
We evaluate methods to visualize human retinal micro-circulation in vivo by standard intensity-based optical coherence
tomography (OCT), speckle-variance optical coherence tomography (svOCT), and phase-variance optical coherence
tomography (pvOCT). En face projection views created from the same volumetric data set of the human retina using all
three data processing methods are created and compared. Additionally we used support vector machine (SVM) based
semi-automatic segmentation to generate en face projection views of individual retinal layers. The layers include: first,
the whole inner retina (from the nerve fiber layer to the outer nuclear layer), and second, from the ganglion cell layer to
the outer nuclear layer. Finally, we compare the retinal vasculature images processed from the three OCT techniques and
fluorescein angiography (FA).
We summarize the performance of an AO-OCT system with reference arm phase shifting for complex conjugate artifactfree
imaging of in vivo retinal structures. As a complex conjugate artifact removal (CCR) method we used a previously
reported technique requiring constant phase shifts between consecutive A-scans. In our system these shifts were
generated by continuous beam path-length changes from offsetting the pivot point of the scanning mirror placed in the
system reference arm. In order to reconstruct the complex spectral fringe pattern we used Fourier transformation along
the transverse axis and a filtering algorithm. The suppression ratio of mirror complex artifact images was assessed based
on acquired in vivo CCR AO-OCT images. Finally, potential problems and limitations connected with this acquisition
scheme and data processing algorithms are discussed.
KEYWORDS: Optical coherence tomography, Visualization, Retina, In vivo imaging, Data acquisition, Phase contrast, Angiography, Imaging systems, Data processing, CMOS sensors
We present high-speed Fourier-domain optical coherence tomography (Fd-OCT) with the phase variance based motion
contrast method for visualizing retinal micro-circulation in vivo. This technique allows non-invasive visualization of a
two-dimensional retinal perfusion map and concurrent volumetric morphology of retinal microvasculature with high
sensitivity. The high-speed acquisition rate at 125kHz A-scans enables reduction of motion artifacts with increased
scanning area if compared to previously reported results. Several scanning schemes with different sampling densities and
scanning areas are evaluated to find optimal parameters for in vivo imaging. In order to evaluate this technique, we
compare OCT micro-capillary imaging using the phase variance technique with fundus fluorescein angiography (FA).
Additionally, volumetric visualization of blood flow for a normal subject is presented.
Single spectrometer-based complex conjugate artifact removal methods are evaluated for in vivo imaging with complementary metal-oxide semiconductor line scan camera based high-speed Fourier-domain optical coherence tomography (FD-OCT) at 100,000 axial scans per second. Performance of three different phase-shifting methods with the same OCT engine is evaluated using modified data acquisition schemes, depending on the requirements of each phase-shifting technique. The suppression ratio of complex conjugate artifact images using a paperboard is assessed for all tested methods. Several other characteristics, including a list of additional hardware requirements (beyond standard FD-OCT components) and data acquisition schemes for each of the methods is presented. In vivo full-range images of human fingerpad and nail are shown and compared with standard FD-OCT images. Additionally, a complex-conjugate-free human retinal volume acquired at the speed of 100,000 A-scans/s is presented.
We describe results of retinal imaging with a novel instrument that combines adaptive optics - Fourier-domain optical
coherence tomography (AO-OCT) with an adaptive optics scanning laser ophthalmoscope (AO-SLO). One of the
benefits of combining Fd-OCT with SLO includes automatic co-registration between the two imaging modalities and the
potential for correcting lateral and transversal eye motion resulting in motion artifact-free volumetric retinal imaging.
Additionally this allows for direct comparison between retinal structures that can be imaged with both modalities (e.g.,
photoreceptor mosaics or microvasculature maps). This dual imaging modality could provide insight into some retinal
properties that could not be accessed by a single imaging system. Additionally, extension of OCT and SLO beyond
structural imaging may open new avenues for diagnostics and testing in ophthalmology. In particular, non-invasive
vasculature mapping with these modalities holds promise of replacing fluorescein angiography in vascular identification.
Several new improvements of our system are described, including results of testing a novel 97-actuator deformable
mirror and AO-SLO light intensity modulation.
We evaluated several, previously published, complex conjugate artifact removal methods and algorithms that have been
proposed for Fourier domain optical coherence tomography (Fd-OCT). To ensure comparable conditions, only one OCT
system was used, but with modified data acquisition schemes, depending on the requirements of each method/algorithm.
This limited our evaluation to single spectrometer based Fd-OCT approaches. The suppression ratio of complex
conjugate artifact images using a paperboard is assessed for all tested methods. Several other metrics are also used for
comparison, including a list of additional hardware requirements (beyond standard Fd-OCT components) and data
acquisition schemes. Finally, in vivo human finger pad and nail images are presented for comparison to the standard Fd-
OCT images and full-range images.
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