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Full-field swept-source optical coherence tomography (FF-SS-OCT) and laser Doppler holography (LDH) are two holographic imaging techniques presenting unique capabilities for ophthalmology. We report on interlaced FF-SS-OCT and LDH imaging with a single instrument. Effectively, retinal blood flow and pulsation could be quasi-simultaneously monitored. This instrument holds potential for a wide scope of ophthalmic applications.

Phase-sensitive optical coherence tomography (OCT) is emerging as an imaging modality that detects functional changes in the retina. Besides imaging photoreceptor function, recently, functional changes in the inner plexiform layer (IPL) have been detected using full-field swept-source OCT. The IPL connects neuronal cells which are dedicated for processing different aspects of the visual information, such as edges in the image or temporal changes. A characteristic of signal processing in the IPL is that different aspects of the visual impression are only processed in very specific depths. Here, we present an investigation of these functional signals for different depths in the IPL with the aim to separate different properties of the visual signal processing. Therefore, we investigate the phase changes of three different sub-layers. Whereas the first two depths, closest to the ganglion cell layer, exhibit an increase in the optical path length, the third depth, closest to the bipolar cell layer, exhibits a decrease in the optical path length. Additionally, we found that the second or middle depth is sensitive to temporal changes, showing a maximum increase of the optical path length at a stimulation frequency of around 10 Hz. The results suggest that the responses from different cell types, which are sensitive to different features of the stimulation signal, can be distinguished by phase-sensitive OCT.

Computational adaptive optics (CAO) is emerging as an attractive alternative to hardware-based solutions for diffraction-limited optical coherence tomography, e.g., of the human retina. Still, to become a reliable and robust solution, many challenges need to be solved. Here, we present CAO based on multiple randomized sub-apertures in combination with suitable filtering to remove disturbing artifacts. We show that this approach can reliably detect aberrations, and we compare results to other algorithms, such as optimization of imaging quality. We also demonstrate that the filtering of reflecting image structures is essential for a robust determination of aberrations.

A method for numerical estimation and correction of aberrations of the eye in fundus imaging with optical coherence tomography (OCT) is presented. Aberrations are determined statistically by using the estimate based on likelihood function maximization. The method can be considered as an extension of the phase gradient autofocusing algorithm in synthetic aperture radar imaging to 2D optical aberration correction. The efficacy of the proposed method has been demonstrated in OCT fundus imaging with 6λ aberrations. After correction, single photoreceptors were resolved. It is also shown that wave front distortions with high spatial frequencies can be determined and corrected.

A method for numerical estimation and correction of aberrations of the eye in fundus imaging with optical coherence tomography (OCT) is presented. Aberrations are determined statistically by using the estimate based on likelihood function maximization. The method can be considered as an extension of the phase gradient autofocusing algorithm in synthetic aperture radar imaging to 2D optical aberrations correction. The efficiency of the proposed method has been demonstrated in OCT fundus imaging with 6λ aberrations. After correction, single photoreceptors were resolved. It is also shown that wavefront distortions with high spatial frequencies can be determined and corrected.Graphical Abstract for Table of Contents[Text. This work is dedicated to development a method for numerical estimation and correction of aberrations of the eye in fundus imaging with OCT. Aberration evaluation is performed statistically by using estimate based on likelihood function maximization. The efficiency of the proposed method has been demonstrated in OCT fundus imaging with 6λ aberrations. It has been shown that spatial high-frequency wavefront distortions can be determined]

We demonstrate functional in vivo imaging of photoreceptor and neuronal layers within the living human retina by looking at the expansion of their optical path length. To this end, we use a special full-field swept-source optical coherence tomography system that acquires all lateral points in parallel, achieving a high-speed data acquisition with up to 200 volumes per second. A combination of computational motion and aberration correction with a suitable phase evaluation scheme yields minuscule changes after exposing the photoreceptors to a white light stimulus.

Non-invasive functional retinal imaging in humans is of tremendous interest. By using phase-sensitive full-field swept-source OCT (FF-SS-OCT) we demonstrated simultaneous quantitative imaging of the optical activation in the photoreceptor and ganglion/inner plexiform layer. Since the signals from the ganglion cells layer are ten-fold smaller than those from the photoreceptor cells a new algorithms for suppression of motion artifacts and pulsatile blood flow in the retinal vessels is important. With improved data evaluation we simultaneously measured the activation of photoreceptors and ganglion/inner plexiform with high quality and were able to analyze the spatial and temporal response of cells in the ganglion/inner plexiform over more than 10 seconds.

Using phase-sensitive full-field swept-source optical coherence tomography we already showed that morphological changes in the photoreceptor outer segments are detectable. Those signals manifest themselves in an elongation of the optical path length. Using improved post.processing we report on progress in detecting signals in the neuronal layers of the human retina. The spatially resolved signals show a characteristic time course and by combining these with simultaneous measurements of the photoreceptors we were able to generate a wiring map of the neuronal retina.

Optische Koharenztomografie (OCT) nutzt Interferenz fur hochaufgeloste Schnitt- oder Volumenbilddarstellungen der Retina. Neue technologische Ansatze haben es in den letzten 25 Jahren immer wieder ermoglicht, die Bildgebungsgeschwindigkeit der OCT zu erhohen. Hierdurch konnten Bewegungsartefakte verringert und Bildfelder vergrosert werden. Die bisher in der Klinik eingesetzten Systeme sind jedoch wegen moglicher Schadigung der Netzhaut durch den fokussierten Strahl und der zur Ablenkung des Strahls notwendigen Mechanik in Geschwindigkeit und raumlicher Prazision der Bildgebung beschrankt. Full-Field-Swept-Source (FF-SS)-OCT parallelisiert die Bildgebung mittels einer Kamera, verzichtet auf bewegte Teile und erlaubt dadurch die Nutzung einer hoheren Lichtleistung. Hier zeigen wir, dass FF-SS-OCT die durch den Herzschlag verursachte Pulsation der Retina mit hoher zeitlicher Auflosung darstellen und vermessen kann. Mit FF-SS-OCT wurden Serien von OCT-Volumen mit 1,8 × 0,7 mm lateraler Ausdehnung uber mehrere Herzzyklen an jungen Probanden aufgenommen. Neben der Morphologie wurden Gefasdarstellungen, der Blutfluss und durch die Druckpulsation ausgeloste Gewebebewegungen gemessen. In den Volumensequenzen waren die wesentlichen Strukturen der neuronalen Retina mit gutem Kontrast und ohne Artefakte durch Augenbewegungen darstellbar. OCT-Angiografie zeigte die retinalen Gefase und Kapillaren. Neben der Morphologie des Gefasnetzwerks wurden anhand von nur einer einzigen FF-SS-OCT-Messung insgesamt 3 dynamische Prozesse des retinalen Gefasnetzwerks untersucht: In den groseren Gefasen konnte die pulsierende Fliesgeschwindigkeit des Blutes mit einer Zeitauflosung im Millisekundenbereich bestimmt werden. Auserdem wurde eine axiale Bewegung der Retina beobachtet, deren zeitlicher Verlauf dem Pulsschlag ahnelt. Diese wird vermutlich durch die Pulsation des Choroids hervorgerufen. Die pulsierenden Druckschwankungen, die sich durch die retinalen Gefase ausbreiten, andern die axiale Ausdehnung der sie umgebenden Retina periodisch um bis zu 2 µm. Hierdurch konnte der Zeitverlauf der Druckanderungen in venosen und arteriellen Gefasen bestimmt werden. Die Verzogerung zwischen der choroidalen und der arteriellen bzw. venosen Pulsation betrug 75 bzw. 110 ms. FF-SS-OCT ermoglicht erstmals die gemeinsame Darstellung dieser schnellen dynamischen Prozesse der Retina.

Aberration-corrected imaging of human photoreceptor cells, whether hardware or software based, presently requires a complex and often expensive setup. Here, we demonstrate a simple and inexpensive off-axis full-field time-domain optical coherence tomography approach to acquire volumetric data of in vivo human retina. Full volumetric, laterally phase stable data are recorded. The lateral phase stability allows computational aberration correction, which enables us to visualize single photoreceptor cells. In addition, our approach is able to correct large aberrations and is thus feasible for the numerical correction of ametropia in post processing. Our implementation of full-field OCT combines a low technical complexity with the possibility to use the phase of the recorded light for computational image correction.

Phase-sensitive coherent imaging exploits changes in the phases of backscattered light to observe tiny alterations of scattering structures or variations of the refractive index. But moving scatterers or a fluctuating refractive index decorrelate the phases and speckle patterns in the images. It is generally believed that once the speckle pattern has changed, the phases are scrambled and any meaningful phase difference to the original pattern is removed. As a consequence, diffusion and tissue motion below the resolution handicap phase-sensitive imaging of biological specimen. Here, we show that, surprisingly, a phase comparison between decorrelated speckle patterns is still possible by utilizing a series of images acquired during decorrelation. The resulting evaluation scheme is mathematically equivalent to methods for astronomic imaging through the turbulent sky by speckle interferometry. We thus adopt the idea of speckle interferometry to phase-sensitive imaging in biological tissues and demonstrate its efficacy for simulated data and for imaging of photoreceptor activity with phase-sensitive optical coherence tomography. The described methods can be applied to any imaging modality that uses phase values for interferometry., Comment: 13 pages, 5 figures

Optical coherence tomography (OCT) has changed diagnostics in ophthalmology by visualizing the morphology of retinal cell layers, which play an important role in human vision. Direct visualization of the activity of these different layers could have an equally strong impact on research and clinical diagnostics. But until recently all attempts to use OCT to image changes of the optical tissue properties that are associated with retinal function (intrinsic optical signals, IOS) were hampered by ocular motion, speckle noise, and limited lateral resolution. Measurements were cumbersome and did not yield straightforward, interpretable physiologic information. Based on a high-speed CMOS camera, full-field swept-source (FF-SS) OCT was developed for in vivo retinal imaging. Within milliseconds, FF-SS-OCT acquires three-dimensional images of the retina which contain information on amplitude and phase of the scattered light virtually free of motion artifacts. Utilizing the phase information, we were able to correct ocular aberrations in order to resolve individual cones and could interferometrically measure sub-wavelength length changes of the photoreceptor outer segments (OS) upon optical stimulation. With this approach, we imaged single cone activity in the living human eye and quantified the photoreceptor response with good accuracy. Currently, the origin of the observed IOS is not completely understood, but there is strong evidence that osmotic effects change the shape of the outer segment. Translating this technology for objective monitoring of photoreceptor activity to clinical applications could have considerable impact in ophthalmology and neurology and might also contribute to basic vision research. This work is an important step to reach the ambitious goal of imaging function of the complete retina with cellular resolution.

Full-field-swept-source optical coherence tomography is capable of detecting small morphological changes in the living human eye below sub-wavelength range by evaluating the phases. This is used to obtain intrinsic optical signals originating in the photoreceptor outer segment, spatially resolved to single photoreceptors. These were measured ex-vivo in explanted porcine retina as well as in the living human eye. The obtained signals are related to an increase of the optical path length of the outer segments. However, they give no hint wether they are caused by an actual physical expansion of the outer segments or by a changes in the index of refraction. Therefore, systematical measurements were carried out to determine the physical nature and biochemical source of the observed effects.

Optical coherence tomography (OCT) images scattering tissues with 5 to 15 μm resolution. This is usually not sufficient for a distinction of cellular and subcellular structures. Increasing axial and lateral resolution and compensation of artifacts caused by dispersion and aberrations is required to achieve cellular and subcellular resolution. This includes defocus which limit the usable depth of field at high lateral resolution. OCT gives access the phase of the scattered light and hence correction of dispersion and aberrations is possible by numerical algorithms. Here we present a unified dispersion/aberration correction which is based on a polynomial parameterization of the phase error and an optimization of the image quality using Shannon’s entropy. For validation, a supercontinuum light sources and a costume-made spectrometer with 400 nm bandwidth were combined with a high NA microscope objective in a setup for tissue and small animal imaging. Using this setup and computation corrections, volumetric imaging at 1.5 μm resolution is possible. Cellular and near cellular resolution is demonstrated in porcine cornea and the drosophila larva, when computational correction of dispersion and aberrations is used. Due to the excellent correction of the used microscope objective, defocus was the main contribution to the aberrations. In addition, higher aberrations caused by the sample itself were successfully corrected. Dispersion and aberrations are closely related artifacts in microscopic OCT imaging. Hence they can be corrected in the same way by optimization of the image quality. This way microscopic resolution is easily achieved in OCT imaging of static biological tissues.

The non-invasive measurement of cellular physiological responses to photostimulation in living retina may have significant clinical value and give new insight into the vision process. Optical coherence tomography (OCT) has been reported to detect suitable intrinsic optical signals (IOS) in retinal photoreceptor layers upon their stimulation. Commonly, changes in backscattering intensity were observed ex vivo and immobilized animals in vivo. However, in humans measurements were time-consuming and cumbersome. Promising results were achieved when observing phase signals to detect intrinsic optical signals. But to achieve sufficient phase stability to image an entire area of photoreceptors turned out to be challenging. Here, we report full-field swept-source OCT to be sufficiently stable to detect the phase signals after projecting a stimulation image onto the living human retina. We extracted time-courses and signal dependencies from the measured datasets. For long stimuli, we were even able to assign responses to single cones. This functional imaging of photoreceptor activity could potentially be used to detect loss of photoreceptor function prior to visible morphological changes, which is associated with numerous retinal diseases.

Time domain OCT measures the interference between sample and reference radiation as a function of the reference arm length. In full-field-OCT (FF-OCT) a camera is used instead of a scanned beam for a parallel detection of the interference pattern and thus acquiring a complete en face image. Because multiple images have to be acquired to resolve the phase ambiguity, this method is prone to motion artifacts. We present a novel motion-insensitive approach to FF-OCT. Spatially coherent illumination and an off-axis reference beam is used to introduce path-length differences between reference and sample light in neighboring pixels. This spatial carrier frequency replaces the temporal carrier frequency in scanned TD-OCT. The setup is based on a Mach-Zehnder interferometer with a super-luminescent diode and a CMOS area camera. The Sensitivity of the system was determined to be 75 dB. The field of view was 1.42 x 1.42 mm. Each frame had 237x237 lateral channels at an axial resolution of 9 µm in tissue. By step-wise changing the length of the reference arm between the en face scans, volumetric in vivo FF-OCT measurements of the human retina have been acquired within 1.3 s. OCT with a spatially coherent off-axis reference beam is suitable for in vivo imaging of human retina. The quality of the images is sufficient to discriminate the different tissue layers.

Functional retinal imaging, especially of neuronal activity non-invasively in humans, is of tremendous interest. Although the activation of photoreceptor cells (PRCs) could be detected in humans, imaging the function of other retinal neurons had been so far hardly possible. Here, using phase-sensitive full-field swept-source optical coherence tomography (FF-SS-OCT), we show simultaneous imaging of the activation in the photoreceptor and ganglion cell layer/inner plexiform layer (GCL/IPL). The signals from the GCL/IPL are 10-fold smaller than those from the PRC and were detectable only using algorithms for suppression of motion artifacts and pulsatile blood flow in the retinal vessels. FF-SS-OCT with improved phase evaluation algorithms, therefore, allowed us to map functional connections between PRC and GCL/IPL, confirming previous ex vivo results. The demonstrated functional imaging of retinal neuronal layers can be a valuable tool in diagnostics and basic research.

Computational adaptive optics (CAO) is emerging as a viable alternative to hardware-based adaptive optics-in particular when applied to optical coherence tomography of the retina. For this technique, algorithms are required that detect wavefront errors precisely and quickly. Here we propose an extension of the frequently used subaperture image correlation. By applying this algorithm iteratively and, more importantly, comparing each subaperture not to the central subaperture but to several randomly selected apertures, we improved aberration correction. Since these modifications only slightly increase the run time of the correction, we believe this method can become the algorithm of choice for many CAO applications.

In optical coherence tomography (OCT), lateral motion is determined either by speckle tracking or by multi-aperture Doppler OCT. Here we show that both methods may provide incorrect results because, outside the focal plane, non-uniform axial motion is misinterpreted as lateral motion. First, we demonstrate the existence of this artifact by means of a simulation for speckle tracking. Then the physical origin of the artifact and its mathematical relation to defocus and axial motion are explained. It is shown that speckle tracking and multi-aperture Doppler OCT are equally affected by the artifact, which has a considerable effect, even for a defocus of less than one Rayleigh length.

With a simple setup, mainly composed of a low coherence light source and a camera, full-field optical coherence tomography (FF-OCT) allows volumetric tissue imaging. However, fringe washout constrains its use in retinal imaging. Here, we present a novel motion-insensitive approach to FF-OCT, which introduces path-length differences between the reference and the sample light in neighboring pixels using an off-axis reference beam. The temporal carrier frequency in scanned time-domain OCT is replaced by a spatial carrier frequency. Volumetric in-vivo FF-OCT measurements of the human retina were acquired in only 1.3 s, comparable to the acquisition times of current clinically used OCT devices.

We demonstrate a new non-invasive method to assess the functional condition of the retinal vascular system. Phase-sensitive full-field swept-source optical coherence tomography (PhS-FF-SS-OCT) is used to investigate retinal vascular dynamics at unprecedented temporal resolution. Motion of retinal tissue, that is induced by expansion of the vessels therein, is measured with an accuracy of about 10 nm. The pulse shape of arterial and venous pulsation, their temporal delay as well as the frequency dependent pulse propagation through the capillary bed are determined. For the first time, imaging speed and motion sensitivity are sufficient for a direct measurement of pulse waves propagating with more than 600 mm/s in retinal vessels of a healthy young subject.

Non-invasive functional imaging of molecular and cellular processes of vision is expected to have immense impact on research and clinical diagnostics. Although suitable intrinsic optical signals (IOS) have been observed ex vivo and in immobilized animals in vivo, it was so far not possible to obtain convincing IOS of photoreceptor activity in humans in vivo. Here, we observed spatially and temporally clearly resolved changes in the optical path length of the photoreceptor outer segment as response to an optical stimulus in living human. To obtain these changes, we evaluated phase data of a parallelized and computationally aberration-corrected optical coherence tomography (OCT) system. The non-invasive detection of optical path length changes shows the neuronal photoreceptor activity of single cones in living human retina, and, more importantly, it provides a new diagnostic option in ophthalmology and neurology and could give new insights into visual phototransduction in humans.

Aberration-corrected imaging of human photoreceptor cells, whether hardware or software based, presently requires a complex and expensive setup. Here we use a simple and inexpensive off-axis full-field time-domain optical coherence tomography (OCT) approach to acquire volumetric data of an in vivo human retina. Full volumetric data are recorded in 1.3 s. After computationally correcting for aberrations, single photoreceptor cells were visualized. In addition, the numerical correction of ametropia is demonstrated. Our implementation of full-field optical coherence tomography combines a low technical complexity with the possibility for computational image correction.

Doppler optical coherence tomography (OCT) quantifies axial motion with high precision, whereas lateral motion cannot be detected by a mere evaluation of phase changes. This problem was solved by the introduction of three-beam Doppler OCT, which, however, entails a high experimental effort. Here, we present the numerical analogue to this experimental approach. Phase-stable complex-valued OCT datasets, recorded with full-field swept-source OCT, are filtered in the Fourier domain to limit imaging to different computational subapertures. These are used to calculate all three components of the motion vector with interferometric precision. As known from conventional Doppler OCT for axial motion only, the achievable accuracy exceeds the actual imaging resolution by orders of magnitude in all three dimensions. The feasibility of this method is first demonstrated by quantifying micro-rotation of a scattering sample. Subsequently, a potential application is explored by recording the 3D motion vector field of tissue during laser photocoagulation in ex-vivo porcine retina.

This paper comments on the article "Retinal pulse wave velocity measurement using spectral-domain optical coherence tomography" by Qian Li et al. The authors propose a method to determine the pulse wave velocity in retinal arteries and veins. This method should enable a noninvasive determination of biomechanical properties of the vessel network, particularly the elasticity of the vessel walls. Although the observations the authors made might seem reasonable at first glance, they are in fact highly surprising and contradictory to theoretical predictions and previously published results.

A full-field speckle interferometry method for non-contact and prospectively high speed Photoacoustic Tomography is introduced and evaluated as proof of concept. Thermoelastic pressure induced changes of the objects topography are acquired in a repetitive mode without any physical contact to the object. In order to obtain high acquisition speed, the object surface is illuminated by laser pulses and imaged onto a high speed camera chip. In a repetitive triple pulse mode, surface displacements can be acquired with nanometre sensitivity and an adjustable sampling rate of e.g. 20 MHz with a total acquisition time far below one second using kHz repetition rate lasers. Due to recurring interferometric referencing, the method is insensitive to thermal drift of the object due to previous pulses or other motion. The size of the investigated area and the spatial and temporal resolution of the detection are scalable. In this study, the approach is validated by measuring a silicone phantom and a porcine skin phantom with embedded silicone absorbers. The reconstruction of the absorbers is presented in 2D and 3D. The sensitivity of the measurement with respect to the photoacoustic detection is discussed. Potentially, Photoacoustic Imaging can be brought a step closer towards non-anaesthetized in vivo imaging and new medical applications not allowing acoustic contact, such as neurosurgical monitoring or burnt skin investigation.

In numerous applications, Fourier-domain optical coherence tomography (FD-OCT) suffers from a limited imaging depth due to signal roll-off, a limited focal range, and autocorrelation noise. Here, we propose a parallel full-field FD-OCT imaging method that uses a swept laser source and an area camera in combination with an off-axis reference, which is incident on the camera at a small angle. As in digital off-axis holography, this angle separates autocorrelation signals and the complex conjugated mirror image from the actual signal in Fourier space. We demonstrate that by reconstructing the signal term only, this approach enables full-range imaging, i.e., it increases the imaging depth by a factor of two, and removes autocorrelation artifacts. The previously demonstrated techniques of inverse scattering and holoscopy can then numerically extend the focal range without loss of lateral resolution or imaging sensitivity. The resulting, significantly enhanced measurement depth is demonstrated by imaging a porcine eye over its entire depth, including cornea, lens, and retina. Finally, the feasibility of in vivo measurements is demonstrated by imaging the living human retina., 12 pages, 8 figures

Full-field swept-source optical coherence tomography (FF-SS-OCT) was recently shown to allow new and exciting applications for imaging the human eye that were previously not possible using current scanning OCT systems. However, especially when using cameras that do not acquire data with hundreds of kHz frame rate, uncorrected phase errors due to axial motion of the eye lead to a drastic loss in image quality of the reconstructed volumes. Here we first give a short overview of recent advances in techniques and applications of parallelized OCT and finally present an iterative and statistical algorithm that estimates and corrects motion-induced phase errors in the FF-SS-OCT data. The presented algorithm is in many aspects adopted from the phase gradient autofocus (PGA) method, which is frequently used in synthetic aperture radar (SAR). Following this approach, the available phase errors can be estimated based on the image information that remains in the data, and no parametrization with few degrees of freedom is required. Consequently, the algorithm is capable of compensating even strong motion artifacts. Efficacy of the algorithm was tested on simulated data with motion containing varying frequency components. We show that even in strongly blurred data, the actual image information remains intact, and the algorithm can identify the phase error and correct it. Furthermore, we use the algorithm to compensate real phase error in FF-SS-OCT imaging of the human retina. Acquisition rates can be reduced by a factor of three (from 60 to 20 kHz frame rate) with an image quality that is even higher compared to uncorrected volumes recorded at the maximum acquisition rate. The presented algorithm for axial motion correction decreases the high requirements on the camera frame rate and thus brings FF-SS-OCT closer to clinical applications.

Phase sensitive OCT enables the measurement of thermal expansion in laser irradiated material at high lateral and temporal resolution. In principle, a calculation of the 3D temperature distribution and its temporal evolution should be possible by evaluating the local expansion. This could be utilized for a non-invasive and very fast temperature measurement, e.g. to realize an online dosimetry for photocoagulation. The possibilities of quantitative investigations at high axial and lateral resolution are demonstrated by imaging the reversible thermal expansion in laser irradiated multilayer silicone phantoms.

We have investigated the magnetic properties of single submicron permalloy rectangles with a thickness of 20 nm and an aspect ratio of 2:1 via anisotropic magnetoresistance AMR . Preparation and investigation via magnetotransport are performed in situ in ultrahigh vacuum. The field-dependent magnetization behavior of the two generic cases with the magnetic field applied perpendicular and parallel to the long axis of the rectangles is studied. Due to the high sensitivity of our setup, single field sweeps are sufficient to obtain magnetoresistance curves of structures with dimensions as small as 600 300 nm2. To link features of the AMR to changes in the micromagnetic states, the remanent state has been investigated via scanning electron microscopy with polarization analysis. Our main result is that the energy density of micromagnetic states can be obtained from the hard-axis magnetization behavior. It is demonstrated that a C/S state can be distinguished from a Landau state and the energy difference between both states is determined.