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Vascular leakage plays a key role in vision-threatening retinal diseases such as diabetic retinopathy and age-related macular degeneration. Fluorescence angiography is the current gold standard for identification of leaky vasculature in vivo, however it lacks depth resolution, providing only 2D images that complicate precise identification and localization of pathological vessels. Optical coherence tomography (OCT) has been widely adopted for clinical ophthalmology due to its high, micron-scale resolution and rapid volumetric scanning capabilities. Nevertheless, OCT cannot currently identify leaky blood vessels. To address this need, we have developed a new method called exogenous contrast-enhanced leakage OCT (ExCEL-OCT) which identifies the diffusion of tracer particles around leaky vasculature following injection of a contrast agent. We apply this method to a mouse model of retinal neovascularization and demonstrate high-resolution 3D vascular leakage measurements in vivo for the first time.

A virtual outreach program was developed specifically for the field of biophotonics. The program consisted of hands-on activities using kits sent to students in advance, real-time video lab tours and prerecorded video content.

Here we describe a new processing pipeline for assessing large amounts of OCT data and apply it to longitudinal imaging data from control and Superoxide Dismutase 1 knockout mice, a model for dry age related macular degeneration. We investigated average axial profiles and compared intensity, spectroscopic, and polarization metrics in the choroid. We found statistically significant differences between groups, suggesting pigmentation changes within the choroid. Analysis of axial profiles reduces the complexity and of analysis which may be valuable in a clinical setting for preventative care, where specific features are not necessarily expected, but trends may be observed over time.

Polarization-sensitive optical coherence tomography (PS-OCT) provides volumetric tissue imaging based on inherent scattering and polarization properties. In fibrous structures such as white matter tissue, PS-OCT enables quantitative mapping of birefringence. For single-input PS-OCT layouts, the accuracy of birefringence measurements strongly depends on the signal-to-noise ratio (SNR) and is prone to mapping artifacts in case of low SNR. Here we present a simple yet effective approach for improving the accuracy of phase retardation and birefringence measurements by performing a noise bias correction of the detected signal amplitudes. We present simulation data to illustrate the influence of the noise bias correction on phase retardation measurements and support our analysis with real-world PS-OCT image data.

Polarization-sensitive optical coherence tomography (PS-OCT) enables three-dimensional imaging of biological tissues based on the inherent contrast provided by scattering and polarization properties. In fibrous tissue such as the white matter of the brain, PS-OCT allows quantitative mapping of tissue birefringence. For the popular PS-OCT layout using a single circular input state, birefringence measurements are based on a straight-forward evaluation of phase retardation data. However, the accuracy of these measurements strongly depends on the signal-to-noise ratio (SNR) and is prone to mapping artifacts when the SNR is low. Here we present a simple yet effective approach for improving the accuracy of PS-OCT phase retardation and birefringence measurements. By performing a noise bias correction of the detected OCT signal amplitudes, the impact of the noise floor on retardation measurements can be markedly reduced. We present simulation data to illustrate the influence of the noise bias correction on phase retardation measurements and support our analysis with real-world PS-OCT image data.

In recent years, there has been growing interest in the application of exogenous contrast agents to supplement the traditional strengths of optical coherence tomography (OCT) and provide additional biological information. In this Letter, we present how indocyanine green, a common fluorescent contrast agent approved by the United States Food and Drug Administration, can provide absorption and spectral contrast for OCT imaging in the mouse eye in vivo. We further demonstrate high stability of spectral contrast measurements for the long-term monitoring of contrast agents in spite of fluctuations in intensity.

In this work, we present a significant step toward in vivo ophthalmic optical coherence tomography and angiography on a photonic integrated chip. The diffraction gratings used in spectral-domain optical coherence tomography can be replaced by photonic integrated circuits comprising an arrayed waveguide grating. Two arrayed waveguide grating designs with 256 channels were tested, which enabled the first chip-based optical coherence tomography and angiography in vivo three-dimensional human retinal measurements. Design 1 supports a bandwidth of 22 nm, with which a sensitivity of up to 91 dB (830 µW) and an axial resolution of 10.7 µm was measured. Design 2 supports a bandwidth of 48 nm, with which a sensitivity of 90 dB (480 µW) and an axial resolution of 6.5 µm was measured. The silicon nitride-based integrated optical waveguides were fabricated with a fully CMOS-compatible process, which allows their monolithic co-integration on top of an optoelectronic silicon chip. As a benchmark for chip-based optical coherence tomography, tomograms generated by a commercially available clinical spectral-domain optical coherence tomography system were compared to those acquired with on-chip gratings. The similarities in the tomograms demonstrate the significant clinical potential for further integration of optical coherence tomography on a chip system., Optical coherence tomography: chip promise The goal of an optical coherence tomography (OCT) imaging system that is integrated on a photonic chip has taken a step closer to reality. Elisabet Rank and coworkers from Austria have shown that arrayed waveguide gratings (AWGs), integrated-optical devices commonly used to separate different wavelength channels in an optical communications system, can be used to replace diffraction gratings in an OCT system. Several designs of silicon nitride AWGs with 256 channels in the near-infrared were fabricated and then tested in an OCT system which was able to capture in-vivo tomograms and angiography of the human eye’s retina, with comparable quality to a conventional system. The next stage of the OCT-on-a-chip research will focus on exploring the use of multimode interference structures, integrated photodiodes, and compact light sources.

In optical coherence tomography (OCT), axial resolution and signal-to-noise ratio (SNR) are typically viewed as uncoupled parameters. We show that this is only true for mirror-like surfaces, and that in diffuse scattering samples such as tissue there is an inherent coupling between axial resolution and measurement SNR. We explain the origin of this coupling and demonstrate that it can be used to achieve increased imaging penetration depth at the expense of resolution. Finally, we argue that this coupling should be considered during OCT system design processes that seek to balance competing needs of resolution, sensitivity, and system/source complexity., Comment: 7 pages, 4 figures

Here we examine sources of spectroscopic contrast, both intrinsic and extrinsic, that can be measured in retinal tissue using optical coherence tomography in a mouse model in vivo with an 840 nm light source.

A post-processing pipeline was created for white light optical coherence tomography, resulting in depth-resolved hyperspectral imaging. This method was tested on both melanin phantoms and in the RPE of rodent eyes, showing that due to its granular nature, melanin can be detected in this manner.

We present a multimodal visible light optical coherence microscopy (OCM) and fluorescence imaging (FI) setup. Specification and phantom measurements were performed to characterize the system. Two applications in neuroimaging were investigated. First, curcumin-stained brain slices of a mouse model of Alzheimer’s disease were examined. Amyloid-beta plaques were identified based on the fluorescence of curcumin, and coregistered morphological images of the brain tissue were provided by the OCM channel. Second, human brain tumor biopsies retrieved intraoperatively were imaged prior to conventional neuropathologic work-up. OCM revealed the three-dimensional structure of the brain parenchyma, and FI added the tumor tissue-specific contrast. Attenuation coefficients computed from the OCM data and the florescence intensity values were analyzed and showed a statistically significant difference for 5-aminolevulinic acid (5-ALA)-positive and -negative brain tissues. OCM findings correlated well with malignant hot spots within brain tumor biopsies upon histopathology. The combination of OCM and FI seems to be a promising optical imaging modality providing complementary contrast for applications in the field of neuroimaging.

Purpose The retinal phenotype of popular mouse models mimicking ophthalmic diseases, such as the superoxide dismutase 1 (SOD1) knockout (KO) mouse model, has mainly been assessed by ex vivo histology and in vivo fundus photography. We used multifunctional optical coherence tomography (OCT) to characterize the retinas of SOD1 KO mice in vivo. Methods The custom-made ophthalmoscope featured a combination of conventional OCT, polarization-sensitive OCT, and OCT angiography. Seven SOD1 KO mice and nine age-matched controls were imaged between 6 and 17 months of age. A postprocessing framework was used to analyze total and outer retinal thickness changes. Drusenlike lesions were segmented, and their sizes and the number of lesions were assessed quantitatively. Their appearance in the conventional reflectivity images, as well as in the corresponding polarization-sensitive images, was characterized qualitatively. Results Drusenlike lesions increased in size and number with age for SOD1 KO mice. Exploiting the multiple contrast channels, the appearance of the lesions was found to resemble pseudodrusen observed in eyes of patients suffering from dry age-related macular degeneration. The total and outer retinal thicknesses were lower on average after 11 months and 7 months in SOD1 KO mice compared with age-matched controls. Neovascularizations were found in one out of seven KO animals. Conclusions OCT imaging proved beneficial for a detailed in vivo characterization of the pathological changes in SOD1 KO mice. Translational Relevance Phenotyping of animal models using modern imaging concepts can be conducted with more precision and might also ease the translation of conclusions between clinical and preclinical research.

We implemented a combined visible light optical coherence microscopy (OCM) and fluorescence imaging platform. A supercontinuum light source in combination with a variable filter box (NKT Photonics) provided a spectral range of 425-680 nm. The OCM setup consists of a Michelson interferometer and a custom made spectrometer. Specification measurements were performed and an axial resolution of 0.88 μm in brain tissue was achieved. The transversal resolution was dependent on the objective lenses and varied from 2 to 8 μm. To change from OCM to fluorescence imaging, two mirrors had to be simply flipped into the light beam in the setup. For acquisition of fluorescence images, a photon multiplier tube (PMT) was used to detect light which had passed through a matched combination of emission, dichroic and excitation filters. As a first proof of concept, a fluorescence phantom consisting of curcumin powder mixed with mounting medium was imaged. The OCM images showed the three-dimensional structure of this phantom and specific contrast was gained by fluorescence imaging. As a control case, mounting medium without curcumin powder was imaged and no fluorescence was observed. One hallmark of Alzheimer's disease (AD) is the development of extracellular amyloid-beta plaques in the brain. The three-dimensional structure of these plaques was investigated with micrometer scale resolution using the OCM system. Curcumin can be used to specifically label amyloid-beta deposits. Curcumin stained brain slices of an AD mouse model were imaged and a specific contrast was gained by the fluorescence.

5-aminolevulinic acid positive brain tumor samples were investigated with a multimodal visible light optical coherence microscope (OCM) and fluorescence imaging (FI) setup. Tumor specific contrast was achieved by the FI part while the three-dimensional tissue morphology was assessed by the OCM part of the setup.

Diagnostic classification techniques used to diagnose cataracts, the world’s leading cause of blindness, are currently based on subjective methods. Here, we present optical coherence tomography as a noninvasive tool for volumetric visualization of lesions formed in the crystalline lens. A custom-made swept-source optical coherence tomography (SS-OCT) system was utilized to investigate the murine crystalline lens. In addition to imaging cataractous lesions in aged wildtype mice, we studied the structure and shape of cataracts in a mouse model of Alzheimer’s disease. Hyperscattering opacifications in the crystalline lens were observed in both groups. Post mortem histological analysis were performed to correlate findings in the anterior and posterior part of the lens to 3D OCT in vivo imaging. Our results showcase the capability of OCT to rapidly visualize cataractous lesions in the murine lens and suggest that OCT might be a valuable tool that provides additional insight for preclinical studies of cataract formation.

We implemented a wide field-of-view visible-light optical coherence microscope (OCM) for investigating ex-vivo brain tissue of patients diagnosed with Alzheimer’s disease (AD) and of a mouse model of AD. A submicrometer axial resolution in tissue was achieved using a broad visible light spectrum. The use of various objective lenses enabled reaching micrometer transversal resolution and the acquisition of images of microscopic brain features, such as cell structures, vessels, and white matter tracts. Amyloid-beta plaques in the range of 10 to [Formula: see text] were visualized. Large field-of-view images of young and old mouse brain sections were imaged using an automated [Formula: see text] stage. The plaque load was characterized, revealing an age-related increase. Human brain tissue affected by cerebral amyloid angiopathy was investigated and hyperscattering structures resembling amyloid beta accumulations in the vessel walls were identified. All results were in good agreement with histology. A comparison of plaque features in both human and mouse brain tissue was performed, revealing an increase in plaque load and a decrease in reflectivity for mouse as compared with human brain tissue. Based on the promising outcome of our experiments, visible light OCM might be a powerful tool for investigating microscopic features in ex-vivo brain tissue.

Polarization-sensitive optical coherence tomography (PS-OCT) provides intrinsic contrast related to tissue microstructure. In the past, PS-OCT has been successfully used for imaging the anterior eye of humans in a variety of pathologic conditions. Here, we present PS-OCT imaging of the anterior eye in mice. Spectral domain PS-OCT centered at a wavelength of 840 nm was performed in anaesthetized laboratory mice. Three dimensional data sets were acquired at a 70 kHz A-line rate. PS-OCT images displaying phase retardation, birefringent axis orientation and degree of polarization uniformity (DOPU) were computed. Similar to human anterior segments, depolarization was observed in the corneal stroma and in structures containing melanin pigments such as the iris and the ciliary body. Birefringence was detected in the sclera close to the limbus. Aside from depolarizing foci observed within structures affected by cataract, the lens appeared mostly polarization preserving. Increased birefringence was observed in a scarred cornea. Given the similarity of the polarization characteristics in the murine eye and the human eye, PS-OCT lends itself as an ideal candidate for non-invasive imaging in preclinical studies in mouse models of anterior segment pathology.

A white light polarization sensitive optical coherence tomography system has been developed, using a supercontinuum laser as the light source. By detecting backscattered light from 400 - 700 nm, an axial resolution of 1.0 µm in air was achieved. The system consists of a free-space interferometer and two homemade spectrometers that detect orthogonal polarization states. Following system specifications, images of a healthy murine retina as acquired by this non-contact system are presented, showing high resolution reflectivity images as well as spectroscopic and polarization sensitive contrast. Additional images of the very-low-density-lipoprotein-receptor (VLDLR) knockout mouse model were acquired. The high resolution allows the detection of small lesions in the retina.

We present three approaches for using optical coherence tomography (OCT) to visualize neuritic plaques in Alzheimer’s disease based on intrinsic optical properties, namely their birefringence, spectroscopic characteristics and scattering profile.

Optical coherence tomography (OCT) is a powerful technology for rapid volumetric imaging in biomedicine. The bright field imaging approach of conventional OCT systems is based on the detection of directly backscattered light, thereby waiving the wealth of information contained in the angular scattering distribution. Here we demonstrate that the unique features of few-mode fibers (FMF) enable simultaneous bright and dark field (BRAD) imaging for OCT. As backscattered light is picked up by the different modes of a FMF depending upon the angular scattering pattern, we obtain access to the directional scattering signatures of different tissues by decoupling illumination and detection paths. We exploit the distinct modal propagation properties of the FMF in concert with the long coherence lengths provided by modern wavelength-swept lasers to achieve multiplexing of the different modal responses into a combined OCT tomogram. We demonstrate BRAD sensing for distinguishing differently sized microparticles and showcase the performance of BRAD-OCT imaging with enhanced contrast for ex vivo tumorous tissue in glioblastoma and neuritic plaques in Alzheimer's disease.

We present a multi-functional optical coherence tomography (OCT) imaging approach to study retinal changes in the very-low-density-lipoprotein-receptor (VLDLR) knockout mouse model with a threefold contrast. In the retinas of VLDLR knockout mice spontaneous retinal-chorodoidal neovascularizations form, having an appearance similar to choroidal and retinal neovascularizations (CNV and RNV) in neovascular age-related macular degeneration (AMD) or retinal angiomatous proliferation (RAP). For this longitudinal study, the mice were imaged every 4 to 6 weeks starting with an age of 4 weeks and following up to the age of 11 months. Significant retinal changes were identified by the multi-functional imaging approach offering a threefold contrast: reflectivity, polarization sensitivity (PS) and motion contrast based OCT angiography (OCTA). By use of this intrinsic contrast, the long-term development of neovascularizations was studied and associated processes, such as the migration of melanin pigments or retinal-choroidal anastomosis, were assessed in vivo. Furthermore, the in vivo imaging results were validated with histological sections at the endpoint of the experiment. Multi-functional OCT proves as a powerful tool for longitudinal retinal studies in preclinical research of ophthalmic diseases. Intrinsic contrast offered by the functional extensions of OCT might help to describe regulative processes in genetic animal models and potentially deepen the understanding of the pathogenesis of retinal diseases such as wet AMD.

The collection and rearing of krill specimens was funded by Australian Antarctic Division science programme Project 4037 (Experimental Krill Biology: Response of krill to environmental change), and Project 4050 (Assessing change in krill distribution and abundance in Eastern Antarctica). MJC is funded by Australian Research Council grant FS110200057. The OCT imaging was funded in part by the U.K. Engineering and Physical Sciences Research Council grant EP/G061688/1, the European Union European Union project FAMOS (FP7 ICT, contract no. 317744) and the CR-UK/EPSRC/MRC/DoH (England) imaging programme. DH was funded by a School of Physics and Astronomy Student Staff Council Vacation Award research studentship. Many small open ocean animals, such as Antarctic krill, are an important part of marine ecosystems. To discover what will happen to animals such as krill in a changing ocean, experiments are run in aquaria where conditions can be controlled to simulate water characteristics predicted to occur in the future. The response of individual animals to changing water conditions can be hard to observe, and with current observation techniques it is very difficult to follow the progress of an individual animal through its life. Optical coherence tomography (OCT) is an optical imaging technique that allows images at high resolution to be obtained from depths up to a few millimeters inside biological specimens. It is compatible with in vivo imaging and can be used repeatedly on the same specimens. In this work, we show how OCT may be applied to post mortem krill samples and how important physiological data such as shell thickness and estimates of organ volume can be obtained. Using OCT we find an average value for the thickness of krill exoskeleton to be (30±4) µm along a 1 cm length of the animal body. We also show that the technique may be used to provide detailed imagery of the internal structure of a pleopod joint and provide an estimate for the heart volume of (0.73±0.03) mm3. Publisher PDF

Maximal safe resection is of utmost importance when operating on intracranial tumors. However, the discrimination between physiological and malignant tissue is challenging. A solution is 5-aminolevulinic acid (5-ALA) induced protoporphyrin IX (PPIX) fluorescence of malignant cells, which has evolved as powerful intraoperative tool for tumor identification. We investigated 43 samples obtained during 5-ALA guided surgery using a combined optical coherence microscopy (OCM) and fluorescence imaging system and compared results to cell density maps utilizing digital pathology. Based on fluorescence intensity, attenuation coefficients, and cell density, tumor core was differentiated from tumor-associated brain parenchyma with high sensitivity and specificity.

Few-mode fibers enable single-shot bright and dark field (BRAD) detection for OCT by collecting the backscattered light into different fiber modes. We demonstrate BRAD imaging with enhanced contrast for characterizing ex vivo tumorous tissue.

A visible light spectral domain optical coherence microscopy system operating in the wavelength range of 450-680 nm was developed. The resulting large wavelength range of 230 nm enabled an ultrahigh axial resolution of 0.88μm in tissue. The setup consisted of a Michelson interferometer combined with a homemade spectrometer with a spectral resolution of 0.03 nm. Scanning of 1 x 1 mm2 and 0.5 x 0.5 mm2 areas was performed by an integrated microelectromechanical mirror. After scanning the light beam is focused onto the tissue by a commercial objective with a 10 x magnification, resulting in a transverse resolution of 2 μm . Specification measurements showed that a -89 dB sensitivity with a 24 dB/mm roll-off could be achieved with the system. First of all the capabilities of the system were tested by investigating millimeter paper, tape and the USAF (US Air Force) 1951 resolution test target. Finally cerebral tissues from non-pathological and Alzheimer's disease affected brains were investigated. The results showed that structures, such as white and gray matter, could be distinguished. Furthermore a first effort was made to differentiate Alzheimer's disease from healthy brain tissue.

We utilized a visible light optical coherence microscopy imaging (OCM) system to investigate the intra-tumor heterogeneity of glioblastoma samples. Using a broad visible spectrum an axial resolution of 0.8 μm was achieved. Objective lenses with short focal lengths were integrated to achieve high transverse resolution. Large field of view images were acquired using an automatic moving x-y-stage. Attenuation maps were calculated from OCM data and cell-density evaluations were performed from corresponding histological sections. Cell density and attenuation data were evaluated. Our analysis revealed intra-tumor heterogeneity by varying cell densities and attenuation coefficients throughout the tumorous areas.

In recent years, there has been growing interest in the application of exogenous contrast agents to supplement the traditional strengths of optical coherence tomography (OCT). Here we present how indocyanine green (ICG), a common fluorescent contrast agent, can provide absorption and spectral contrast for OCT imaging in the mouse eye in vivo. We further demonstrate high stability of spectral contrast measurements for long term monitoring of contrast agents despite intensity fluctuations. This type of contrast-enhanced OCT may be able to provide similar information to ICG fluorescence angiography, but with the added benefit of depth-resolution and perfectly coregistered OCT intensity data.

A polarization sensitive extension was implemented into our bright and dark field optical coherence tomography (BRAD-OCT) system. The few-mode fiber detection enables the analysis of the scattering profiles of biological tissues by sensing the sample backscattering information at larger angles through the different fiber modes, enabling simultaneous detection of the bright and dark field. Since the polarization state of light scattered at different angles strongly depends on the tissue microstructure, the polarization sensitive detection improves the imaging capabilities of BRAD-OCT. The system performance was evaluated in phantoms of differently sized microparticles, showing different polarization characteristics for the different fiber modes. Preliminary analysis has been performed in ex vivo brain tissue, where stokes vectors analysis of the different modal images reconstructed by our BRAD-OCT setup indicates that polarization sensitive in combination with BRAD detection is a promising tool for investigating the scattering properties of biological tissues.

A multimode fiber detection for angle-resolved optical coherence tomography is presented. A relation between the angle of reflection from the illuminated sample and the intensity distribution of the modes into a multimode fiber was demonstrated.

A spectral domain optical coherence microscopy system operating in the visible wavelength range was developed. With an axial resolution of 0.9 μm cerebral tissues from non-pathological and Alzheimer’s disease affected brains were investigated.

A few-mode fiber based detection for OCT systems is presented. The capability of few-mode fibers for delivering light through different fiber paths enables the application of these fibers for angular scattering tissue characterization. Since the optical path lengths traveled in the fiber change between the fiber modes, the OCT image information will be reconstructed at different depth positions, separating the directly backscattered light from the light scattered at other angles. Using the proposed method, the relation between the angle of reflection from the sample and the respective modal intensity distribution was investigated. The system was demonstrated for imaging ex-vivo brain tissue samples of patients with Alzheimer’s disease.

We present a polarization-sensitive (PS) extension for bright- and dark-field (BRAD) optical coherence tomography imaging. Using a few-mode fiber detection scheme, the light backscattered at different angles is separated, and the BRAD images of tissue scattering are generated. A calibration method to correct for the fiber birefringence is proposed. Since particle scattering profiles are polarization dependent, a PS detection extends the capabilities for investigating the scattering properties of biological tissues. Both phantoms consisting of different-sized microparticles and a brain tissue specimen were imaged to validate the system performance and demonstrate the complementary image contrast.

Significance. Recent Alzheimer’s disease (AD) patient studies have focused on retinal analysis, as the retina is the only part of the central nervous system that can be imaged noninvasively by optical methods. However, as this is a relatively new approach, the occurrence and role of retinal pathological features are still debated. Aim. The retina of an APP/PS1 mouse model was investigated using multicontrast optical coherence tomography (OCT) in order to provide a documentation of what was observed in both transgenic and wild-type mice. Approach. Both eyes of 24 APP/PS1 transgenic mice (age: 45 to 104 weeks) and 15 age-matched wild-type littermates were imaged by the custom-built OCT system. At the end of the experiment, retinas and brains were harvested from a subset of the mice (14 transgenic, 7 age-matched control) in order to compare the in vivo results to histological analysis and to quantify the cortical amyloid beta plaque load. Results. The system provided a combination of standard reflectivity data, polarization-sensitive data, and OCT angiograms. Qualitative and quantitative information from the resultant OCT images was extracted on retinal layer thickness and structure, presence of hyper-reflective foci, phase retardation abnormalities, and retinal vasculature. Conclusions. Although multicontrast OCT revealed abnormal structural properties and phase retardation signals in the retina of this APP/PS1 mouse model, the observations were very similar in transgenic and control mice.