(In Reverse Order of publication, most recent first)
Title: Fast visible and extended near-infrared multispectral fundus camera
Authors:T. Alterini, F. Diaz, F. J. Burgos-Fernandez, L. Gonzalez, C. Mateo and M. Vilaseca
Abstract: We present a multispectral fundus camera that performs fast imaging of the ocular posterior pole in the visible and near-infrared (400 to 1300 nm) wavelengths through 15 spectral bands, using a flashlight source made of light-emitting diodes, and CMOS and InGaAs cameras. We investigate the potential of this system for visualizing occult and overlapping structures of the retina in the unexplored wavelength range beyond 900 nm, in which radiation can penetrate deeper into the tissue. Reflectance values at each pixel are also retrieved from the acquired images in the analyzed spectral range. The available spectroscopic information and the visualization of retinal structures, specifically the choroidal vasculature and drusen-induced retinal pigment epithelium degeneration, which are hardly visible in conventional color fundus images, underline the clinical potential of this system as a new tool for ophthalmic diagnosis.
Title: Experimental characterization of the speckle pattern at the output of a multimode optical fiber
Authors:Donatus Halpaap, Jordi Tiana-Alsina, Meritxell Vilaseca, and Cristina Masoller
Abstract: Speckle patterns produced by coherent waves interfering with each other are undesirable in many imaging applications (for example, in laser projection systems) but on the other hand, they contain useful information that can be exploited (for example, for blood flow analysis or reconstruction of the object that generates the speckle). It is therefore important to understand how speckle can be enhanced or reduced by tailoring the coherence of laser light. Using a conventional semiconductor laser and a multimode optical fiber we study experimentally how the speckle pattern depends on the laser pump current and on the image acquisition settings. By varying the pump current from below to above the lasing threshold, and simultaneously tuning the image exposure time to compensate for the change in brightness, we find conditions that allow for recorded images with similar average intensity, but with speckle contrast (the standard deviation of the intensity over the average intensity) as low as 0.16, or as high as 0.99.
Title: Network-based features for retinal fundus vessel structure analysis
Authors: Pablo Amil, Cesar F Reyes-Manzano, Lev Guzmán-Vargas, Irene Sendiña-Nadal and Cristina Masoller
Abstract: Retinal fundus imaging is a non-invasive method that allows visualizing the structure of the blood vessels in the retina whose features may indicate the presence of diseases such as diabetic retinopathy (DR) and glaucoma. Here we present a novel method to analyze and quantify changes in the retinal blood vessel structure in patients diagnosed with glaucoma or with DR. First, we use an automatic unsupervised segmentation algorithm to extract a tree-like graph from the retina blood vessel structure. The nodes of the graph represent branching (bifurcation) points and endpoints, while the links represent vessel segments that connect the nodes. Then, we quantify structural differences between the graphs extracted from the groups of healthy and non-healthy patients. We also use fractal analysis to characterize the extracted graphs. Applying these techniques to three retina fundus image databases we find significant differences between the healthy and non-healthy groups (p-values lower than 0.005 or 0.001 depending on the method and on the database). The results are sensitive to the segmentation method (manual or automatic) and to the resolution of the images.
Title: High-speed OCT-based ocular biometer combined with an air-puff system for determination of induced retraction-free eye dynamics
Authors: Alfonso Jiménez-villar, Ewa Mączyńska, Artur Cichański, Maciej Wojtkowski, Bartłomiej J. Kałużny, and Ireneusz Grulkowski
Abstract: We demonstrate a swept source OCT-based ocular biometer integrated with an air-puff stimulus to study the reaction of the eye to mechanical stimulation in vivo. The system enables simultaneous measurement of the stimulus strength and high-speed imaging of the eye dynamics along the visual axis. We characterize the stimulus and perform optimization of the data acquisition for a proper interpretation of the results. Access to the dynamics of axial eye length allows for a determination of the eye retraction, which is used to correct the air-puff induced displacement of ocular structures. We define the parameters to quantify the reaction of the eye to the air puff and determine their reproducibility in a group of healthy subjects. We observe the corneal deformation process and axial wobbling of the crystalline lens. OCT biometer combined with the air puff is the first instrument with the potential to provide comprehensive information on the biomechanics of ocular components.
Title: Air-Puff-Induced Dynamics of Ocular Components Measured with Optical Biometry
Authors: Ewa Maczynska, Jagoda Rzeszewska-Zamiara, Alfonso Jimenez Villar, Maciej Wojtkowski, Bartlomiej J. Kaluzny, and Ireneusz Grulkowski
Abstract: To analyze the dynamics of all optical components of the eye and the behavior of the eyeball under air-puff conditions in vivo. To determine the impact of the intraocular pressure (IOP) on the air-puff-induced deformation of the eye. Twenty eyes of 20 healthy subjects were included in this study. The dynamics of the ocular components, such as the cornea, the crystalline lens, and the retina, was measured by a prototype swept source optical coherence tomography biometer integrated with the air-puff system. The system allows to acquire a series of axial scans at the same location as a function of time with no transverse scanning. Several parameters were extracted from optical coherence tomography data. The IOP was measured using a Goldmann applanation tonometry. The measurements of the eyes were performed before and 2 hours after administration of IOP-reducing drops, namely, 0.2 % brimonidine tartrate.There is a statistically significant correlation of corneal thickness, vitreous depth, and eye length with IOP. The deformation amplitudes of the cornea and the crystalline lens are inversely proportional to the IOP, but statistical significance is achieved only for the cornea. The crystalline lens is displaced without compression, and the return has the form of wobbling. The reduction of IOP level induces corresponding changes in the extracted parameters. Optical biometry combined with air puff provides comprehensive information on the in vivo behavior of all ocular components, including the crystalline lens. Measurement of the axial length dynamics of during deformation enables correcting the deformation for eye retraction.
Title: Spatiotemporal optical coherence (STOC) manipulation suppresses coherent cross-talk in full-field swept-source optical coherence tomography
Authors: Dawid Borycki, Michał Hamkało, Maciej Nowakowski, Maciej Szkulmowski, and Maciej Wojtkowski
Abstract: Full-field swept-source optical coherence tomography (FF-SS-OCT) provides high-resolution depth-resolved images of the sample by parallel Fourier-domain interferometric detection. Although FF-SS-OCT implements high-speed volumetric imaging, it suffers from the cross-talk-generated noise from spatially coherent lasers. This noise reduces the transversal image resolution, which in turn, limits the wide adaptation of FF-SS-OCT for practical and clinical applications. Here, we introduce the novel spatiotemporal optical coherence (STOC) manipulation. In STOC the time-varying inhomogeneous phase masks are used to modulate the light incident on the sample. By properly adjusting these phase masks, the spatial coherence can be reduced. Consequently, the cross-talk-generated noise is suppressed, the transversal image resolution is improved by the factor of2−−√, and sample features become visible. STOC approach is validated by imaging 1951 USAF resolution test chart covered by the diffuser, scattering phantom and the rat skin ex vivo. In all these cases STOC suppresses the cross-talk-generated noise, and importantly, do not compromise the transversal resolution. Thus, our method provides an enhancement of FF-SS-OCT that can be beneficial for imaging biological samples.
Title: Speckle reduction in double-pass retinal images
Authors: Donatus Halpaap, Carlos E. García-Guerra, Meritxell Vilaseca and Cristina Masoller
Abstract: The double pass (DP) technique quantifies the optical quality of the eye by measuring its point spread function. The low reflectivity of the retina requires the use of a high-brightness, point-like illumination source, and thus, DP systems use laser diodes (LDs). However, LDs light produces speckle, and a low-cost solution to reduce speckle is to include a vibrating mirror in the beam path. With the goal of finding an all-optical solution, here we perform a comparative study of the amount of speckle produced by three semiconductor light sources: an LD, a light emitting diode (LED), and a superluminescent diode (SLED). We also compare the results with the speckle reduction that is obtained with a vibrating mirror. We find that the SLED is a good alternative to LD illumination, as the amount of speckle in the image is almost as low as that obtained with an LD and a vibrating mirror in the beam path.
Title: Unsupervised feature extraction of anterior chamber OCT images for ordering and classification
Authors: Pablo Amil, Laura González, Elena Arrondo, Cecilia Salinas, Josep Lluís Guell, Cristina Masoller and Ulrich Parlitz
Abstract: We propose an image processing method for ordering anterior chamber optical coherence tomography (OCT) images in a fully unsupervised manner. The method consists of three steps: Firstly we preprocess the images (filtering the noise, aligning and normalizing the resolution); secondly, a distance measure between images is computed for every pair of images; thirdly we apply a machine learning algorithm that exploits the distance measure to order the images in a two-dimensional plane. The method is applied to a large (~1000) database of anterior chamber OCT images of healthy subjects and patients with angle-closure and the resulting unsupervised ordering and classification is validated by two ophthalmologists.
Title: Energy-Reduced Arrhythmia Termination Using Global Photostimulation in Optogenetic Murine Hearts
Authors: Raúl A. Quiñonez Uribe, Stefan Luther, Laura Diaz-Maue and Claudia Richter
Abstract: Complex spatiotemporal non-linearity as observed during cardiac arrhythmia strongly correlates with vortex-like excitation wavelengths and tissue characteristics. Therefore, the control of arrhythmic patterns requires fundamental understanding of dependencies between onset and perpetuation of arrhythmia and substrate instabilities. Available treatments, such as drug application or high-energy electrical shocks, are discussed for potential side effects resulting in prognosis worsening due to the lack of specificity and spatiotemporal precision. In contrast, cardiac optogenetics relies on light sensitive ion channels stimulated to trigger excitation of cardiomyocytes solely making use of the inner cell mechanisms. This enables low-energy, non-damaging optical control of cardiac excitation with high resolution. Recently, the capability of optogenetic cardioversion was shown in Channelrhodopsin-2 (ChR2) transgenic mice. But these studies used mainly structured and local illumination for cardiac stimulation. In addition, since optogenetic and electrical stimulus work on different principles to control the electrical activity of cardiac tissue, a better understanding of the phenomena behind optogenetic cardioversion is still needed. The present study aims to investigate global illumination with regard to parameter characterization and its potential for cardioversion. Our results show that by tuning the light intensity without exceeding 1.10 mW mm-2, a single pulse in the range of 10–1,000 ms is sufficient to reliably reset the heart into sinus rhythm. The combination of our panoramic low-intensity photostimulation with optical mapping techniques visualized wave collision resulting in annihilation as well as propagation perturbations as mechanisms leading to optogenetic cardioversion, which seem to base on other processes than electrical defibrillation. This study contributes to the understanding of the roles played by epicardial illumination, pulse duration and light intensity in optogenetic cardioversion, which are the main variables influencing cardiac optogenetic control, highlighting the advantages and insights of global stimulation. Therefore, the presented results can be modules in the design of novel illumination technologies with specific energy requirements on the way toward tissue-protective defibrillation techniques.
Title: Three-photon light-sheet fluorescence microscopy
Authors: A. Escobet-Montalbán, F. M. Gasparoli, J. Nylk, P. Liu, Z. Yang, and K. Dholakia
Abstract: We present the first demonstration of three-photon excitation light-sheet fluorescence microscopy. Light-sheet fluorescence microscopy in single- and two-photon modes has emerged as a powerful wide-field, low-photodamage technique for fast volumetric imaging of biological samples. We extend this imaging modality to the three-photon regime, enhancing its penetration depth. Our present study uses a conventional femtosecond pulsed laser at 1000 nm wavelength for the imaging of 450 μm diameter cellular spheroids. In addition, we show, experimentally and through numerical simulations, the potential advantages in three-photon light-sheet microscopy of using propagation-invariant Bessel beams in preference to Gaussian beams.
Title: Wide-field multiphoton imaging through scattering media without correction.
Authors: A. Escobet-Montalbán, R. Spesyvtsev, M. Chen, W. Afshar Saber, M. Andrews, C. Simon Herrington, M. Mazilu and K. Dholakia
Abstract: Optical approaches to fluorescent, spectroscopic, and morphological imaging have made exceptional advances in the last decade. Super-resolution imaging and wide-field multiphoton imaging are now underpinning major advances across the biomedical sciences. While the advances have been startling, the key unmet challenge to date in all forms of optical imaging is to penetrate deeper. A number of schemes implement aberration correction or the use of complex photonics to address this need. In contrast, we approach this challenge by implementing a scheme that requires no a priori information about the medium nor its properties. Exploiting temporal focusing and single-pixel detection in our innovative scheme, we obtain wide-field two-photon images through various turbid media including a scattering phantom and tissue reaching a depth of up to seven scattering mean free path lengths. Our results show that it competes favorably with standard point-scanning two-photon imaging, with up to a fivefold improvement in signal-to-background ratio while showing significantly lower photobleaching.
Be-Optical Conference Proceedings
(In Reverse Order of publication, most recent first)
Proceedings of SPIE 11073, 110730Y (2019).
Title: Hyperspectral eye fundus imaging with extended spectral range towards the near infrared
Authors: T. Alterini, F. Díaz-Doutón, M. Vilaseca
Abstract: Eye fundus photography routinely used in clinical practice is restricted to color imaging of the retina. In the last years, hyperspectral imaging has shown to be a powerful tool for the spectral analysis of biological tissue. In this study, we present a fully custom-made fast hyperspectral fundus camera based on light emitting diodes (LED) with 15 different wavelengths of emission and with extended spectral sensitivity towards the near infrared (NIR) (from 400 nm to 1300 nm), which allows imaging deeper retinal layers, including the choroid, than current clinical devices. These new features will be very useful for a better understanding of ocular diseases as well as aiding in their diagnosis.
Title: Characterization of speckle patterns generated by a semiconductor laser with optical feedback for speckle reduction in retinal imaging instruments
Authors: D. Halpaap, J. Tiana-Alsina, M. Vilaseca, C. Masoller
Abstract: We study experimentally the operating conditions of a semiconductor laser diode subjected to different amounts of optical feedback in order to find a stable and cost-efficient solution for speckle reduction in double-pass retinal imaging.
Title: Optimization of a SS-OCT with a focus tunable lens for enhanced visualization of ocular opacities
Authors: A. Rodriguez-Aramendia, I. Grulkowski, A. Jimenez-Villar, S. Manzanera, Y. Chen, J. Mompean, F. Díaz-Doutón, J. Pujol, J. L. Güell and P. Artal
Abstract: Optical methods have been recently used to perform objective assessment of crystalline lens and corneal opacities. Swept-source optical coherence tomography (SS-OCT) enables measurements of the back-reflected or back-scattered photons from the internal objects. In this work, we present a long-depth range SS-OCT system, with a focus tunable lens, optimized for the visualization of large sections of the posterior segment of the eye, including the vitreous. The system was validated using an eye model.
Title: Wide-field multiphoton imaging with TRAFIX
Authors: A. Escobet-Montalbán, P. Wijesinghe, M. Chen and K. Dholakia
Abstract: Optical approaches have broadened their impact in recent years with innovations in both wide-field and super- resolution imaging, which now underpin biological and medical sciences. Whilst these advances have been remarkable, to date, the ongoing challenge in optical imaging is to penetrate deeper. TRAFIX is an innovative approach that combines temporal focusing illumination with single-pixel detection to obtain wide-field multi- photon images of fluorescent microscopic samples deep through scattering media without correction. It has been shown that it can image through biological samples such as rat brain or human colon tissue up to a depth of seven scattering mean-free-path lengths. Comparisons of TRAFIX with standard point-scanning two-photon microscopy show that the former can yield a five-fold higher signal-to-background ratio while significantly reducing photobleaching of the specimen. Here, we show the first preliminary demonstration of TRAFIX with three-photon excitation imaging dielectric beads. We discuss the advantages of the TRAFIX approach combined with compressive sensing for biomedicine.
Title: Imaging Through Turbid Media with Wavefront Modulated Illumination Full-Field Optical Coherence Microscopy
Authors: M. Hamkalo, D. Borycki and M. Wojtkowski
Abstract: Imaging deep inside the tissue still remains a challenge for all microscopic techniques. Imaging using coherent illumination is even more challenging due to unwanted effects like speckle formation causing significant loss of imaging contrast. In our work we propose a solution to this problem by controlling the spatial distribution of phase of light illuminating the sample. In newly developed optical set-up the beam illuminating the sample first is passing through SLM that enables to fully control projected light patterns. The interferometric setup enables to perform 3D full field OCM imaging. We present the results from controlled wavefront illumination and its ability to image through scattering layers. The ultimate goal of our techniques is to create new imaging method applicable for biological 3D imaging in turbid medium.
Title: Incoherent Light Sources for Speckle Reduction in Double Pass Ocular Imaging
Authors: D. Halpaap, M. Vilaseca and C. Masoller
Abstract: The double pass imaging method is used to obtain the point spread function of a patient’s eye; however it suffers from speckle formation. Here we present a comparison of speckle formation in double pass imaging using three different semiconductor-based light sources.