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Optical coherence tomography (OCT) has seen widespread success as an in vivo clinical diagnostic 3D imaging modality, impacting areas including ophthalmology, cardiology, and gastroenterology. Despite its many advantages, such as high sensitivity, speed, and depth penetration, OCT suffers from several shortcomings that ultimately limit its utility as a 3D microscopy tool, such as its pervasive coherent speckle noise and poor lateral resolution required to maintain millimeter-scale imaging depths. Here, we present 3D optical coherence refraction tomography (OCRT), a computational extension of OCT which synthesizes an incoherent contrast mechanism by combining multiple OCT volumes, acquired across two rotation axes, to form a resolution-enhanced, speckle-reduced, refraction-corrected 3D reconstruction. Our label-free computational 3D microscope features a novel optical design incorporating a parabolic mirror to enable the capture of 5D plenoptic datasets, consisting of millimetric 3D fields of view over up to $\pm75^\circ$ without moving the sample. We demonstrate that 3D OCRT reveals 3D features unobserved by conventional OCT in fruit fly, zebrafish, and mouse samples.

Illuminating or imaging samples from a broad angular range is essential in a wide variety of computational 3D imaging and resolution-enhancement techniques, such as optical projection tomography (OPT), optical diffraction tomography (ODT), synthetic aperture microscopy, Fourier ptychographic microscopy (FPM), structured illumination microscopy (SIM), photogrammetry, and optical coherence refraction tomography (OCRT). The wider the angular coverage, the better the resolution enhancement or 3D resolving capabilities. However, achieving such angular ranges is a practical challenge, especially when approaching plus-or-minus 90 degrees or beyond. Often, researchers resort to expensive, proprietary high numerical aperture (NA) objectives, or to rotating the sample or source-detector pair, which sacrifices temporal resolution or perturbs the sample. Here, we propose several new strategies for multi-angle imaging approaching 4pi steradians using concave parabolic or ellipsoidal mirrors and fast, low rotational inertia scanners, such as galvanometers. We derive theoretically and empirically relations between a variety of system parameters (e.g., NA, wavelength, focal length, telecentricity) and achievable fields of view (FOVs) and importantly show that intrinsic tilt aberrations do not restrict FOV for many multi-view imaging applications, contrary to conventional wisdom. Finally, we present strategies for avoiding spherical aberrations at obliquely illuminated flat boundaries. Our simple designs allow for high-speed multi-angle imaging for microscopic, mesoscopic, and macroscopic applications.

Anterior uveitis is the most common form of intraocular inflammation, and one of its main signs is the presence of white blood cells (WBCs) in the anterior chamber (AC). Clinically, the true composition of cells can currently only be obtained using AC paracentesis, an invasive procedure to obtain AC fluid requiring needle insertion into the AC. We previously developed a spectroscopic optical coherence tomography (SOCT) analysis method to differentiate between populations of RBCs and subtypes of WBCs, including granulocytes, lymphocytes and monocytes, both in vitro and in ACs of excised porcine eyes. We have shown that different types of WBCs have distinct characteristic size distributions, extracted from the backscattered reflectance spectrum of individual cells using Mie theory. Here, we further develop our method to estimate the composition of blood cell mixtures, both in vitro and in vivo. To do so, we estimate the size distribution of unknown cell mixtures by fitting the distribution observed using SOCT with a weighted combination of reference size distributions of each WBC type calculated using kernel density estimation. We validate the accuracy of our estimation in an in vitro study, by comparing our results for a given WBC sample mixture with the cellular concentrations measured by a hemocytometer and SOCT images before mixing. We also conducted a small in vivo quantitative cell mixture validation pilot study which demonstrates congruence between our method and AC paracentesis in two patients with uveitis. The SOCT based method appears promising to provide quantitative diagnostic information of cellular responses in the ACs of patients with uveitis.

We present a feature-free photogrammetric technique that enables quantitative 3D mesoscopic (mm-scale height variation) imaging with tens-of-micron accuracy from sequences of images acquired by a smartphone at close range (several cm) under freehand motion without additional hardware. Our end-to-end, pixel-intensity-based approach jointly registers and stitches all the images by estimating a coaligned height map, which acts as a pixel-wise radial deformation field that orthorectifies each camera image to allow homographic registration. The height maps themselves are reparameterized as the output of an untrained encoder-decoder convolutional neural network (CNN) with the raw camera images as the input, which effectively removes many reconstruction artifacts. Our method also jointly estimates both the camera's dynamic 6D pose and its distortion using a nonparametric model, the latter of which is especially important in mesoscopic applications when using cameras not designed for imaging at short working distances, such as smartphone cameras. We also propose strategies for reducing computation time and memory, applicable to other multi-frame registration problems. Finally, we demonstrate our method using sequences of multi-megapixel images captured by an unstabilized smartphone on a variety of samples (e.g., painting brushstrokes, circuit board, seeds).

We present a general theory of optical coherence tomography (OCT), which synthesizes the fundamental concepts and implementations of OCT under a common 3D k-space framework. At the heart of this analysis is the Fourier diffraction theorem, which relates the coherent interaction between a sample and plane wave to the Ewald sphere in the 3D k-space representation of the sample. While only the axial dimension of OCT is typically analyzed in k-space, we show that embracing a fully 3D k-space formalism allows explanation of nearly every fundamental physical phenomenon or property of OCT, including contrast mechanism, resolution, dispersion, aberration, limited depth of focus, and speckle. The theory also unifies diffraction tomography, confocal microscopy, point-scanning OCT, line-field OCT, full-field OCT, Bessel-beam OCT, transillumination OCT, interferometric synthetic aperture microscopy (ISAM), and optical coherence refraction tomography (OCRT), among others. Our unified theory not only enables clear understanding of existing techniques, but also suggests new research directions to continue advancing the field of OCT.

Recently, there has been growing interest and effort in developing high-speed high-precision 3D imaging technologies for a wide range of industrial, automotive and biomedical applications. Optical frequency-modulated continuous wave (FMCW) light detection and ranging (LiDAR), which shares the same working principle as swept-source optical coherence tomography (SSOCT), is an emerging 3D surface imaging technology that offers higher sensitivity and resolution than conventional time-of-flight (ToF) ranging. Recently, with the development of high-performance swept sources with meter-scale instantaneous coherence lengths, the imaging range of both OCT and FMCW has been significantly improved. However, due to the limited bandwidth of current generation digitizers and the speed limitations of beam steering using mechanical scanners, long range OCT and FMCW typically suffer from a low 3D frame rate (<1Hz), which greatly restricts their applications in imaging dynamic or moving objects. In this work, we report a high-speed FMCW based 3D surface imaging system, combining a grating for beam steering with a compressed time-frequency analysis approach for depth retrieval. We thoroughly investigate the localization accuracy and precision of our system both theoretically and experimentally. Finally, we demonstrate 3D surface imaging results of multiple static and moving objects, including a flexing human hand. The demonstrated technique performs 3D surface imaging with submillimeter localization accuracy over a tens-of-centimeter imaging range with an overall depth voxel acquisition rate of 7.6 MHz, enabling densely sampled 3D surface imaging at video rate.

Robotic automation has the potential to assist human surgeons in performing suturing tasks in microsurgery, and in order to do so a robot must be able to guide a needle with sub-millimeter precision through soft tissue. This paper presents a robotic suturing system that uses 3D optical coherence tomography (OCT) system for imaging feedback. Calibration of the robot-OCT and robot-needle transforms, wound detection, keypoint identification, and path planning are all performed automatically. The calibration method handles pose uncertainty when the needle is grasped using a variant of iterative closest points. The path planner uses the identified wound shape to calculate needle entry and exit points to yield an evenly-matched wound shape after closure. Experiments on tissue phantoms and animal tissue demonstrate that the system can pass a suture needle through wounds with 0.27 mm overall accuracy in achieving the planned entry and exit points., Comment: 7 pages, 11 figures, 2 tables. Accepted to the 2020 International Conference on Robotics and Automation (ICRA)

We report a novel extension to structured illumination (SI) microscopy that utilizes the Talbot self-imaging effect to generate a SI pattern on a sample with field-of-view (FOV) and resolution unconstrained by the numerical aperture (NA) of the microscope objective. This enables high-resolution SI pattern generation (up to the free-space propagation limit) across large FOVs, which in turn allows SI microscopy to be efficiently adapted for high-throughput imaging. We demonstrated this concept using a simple prototype microscope to image a large FOV (450 x 350 um) with a diffraction-limited fluorescence resolution limited by the microscope NA to 2 um. A custom-fabricated structured element, designed with periodic spatial features beyond the diffraction limit of the microscope imaging objective, was positioned to within an integer multiple of half the Talbot distance away from the sample. This generated a large FOV, sub-diffraction SI pattern at the sample plane via the Talbot self-imaging effect. Conventional SI super-resolution principles were then used to increase imaging resolution to 500 nm across the complete FOV, roughly a factor of 4 improvement over the microscope NA diffraction limit. This approach should also be applicable for high-throughput sub-diffraction resolution multimodal imaging., Comment: 10 pages, 3 figures

Though structured illumination (SI) microscopy is a popular imaging technique conventionally associated with fluorescent super-resolution, recent works have suggested its applicability towards sub-diffraction coherent imaging with quantitative endogenous biological contrast. Here, we demonstrate that SI can efficiently integrate the principles of fluorescent super-resolution and coherent synthetic aperture to achieve 3D dual-modality sub-diffraction resolution, fluorescence and refractive-index (RI) visualizations of biological samples. Such a demonstration can enable future biological studies to synergistically explore molecular and biophysical/biochemical mechanisms at sub-diffraction resolutions. We experimentally demonstrate this framework by introducing a SI microscope capable of 3D sub-diffraction fluorescence and RI imaging, and verify its biological visualization capabilities by experimentally reconstructing 3D RI/fluorescence visualizations of fluorescent calibration microspheres as well as alveolar basal epithelial adenocarcinoma (A549) and human colorectal adenocarcinmoa (HT-29) cells, fluorescently stained for F-actin., Comment: 24 pages, 6 figures in main text, 3 figures in supplementary text

This work introduces a novel reinterpretation of structured illumination (SI) microscopy for coherent imaging that allows three-dimensional imaging of complex refractive index (RI). To do so, we show that coherent SI is mathematically equivalent to a superposition of angled illuminations. It follows that raw acquisitions for standard SI-enhanced quantitative-phase images can be processed into complex electric-field maps describing sample diffraction under angled illuminations. Standard diffraction tomography (DT) computation can then be used to reconstruct the sample 3D RI distribution at sub-diffraction resolutions. We demonstrate this concept by using a SI-quantitative-phase imaging system to computationally reconstruct 3D RI distributions of human breast (MCF-7) and colorectal (HT-29) adenocarcinoma cells. Our experimental setup uses a spatial light modulator to generate structured patterns at the sample and collects angle-dependent sample diffraction using a common-path, off-axis interference configuration with no moving components. Furthermore, this technique holds promise for easy pairing with SI fluorescence microscopy, and important future extensions may include multimodal, sub-diffraction resolution, 3D RI and fluorescent visualizations., Comment: 19 total pages (including references), 8 figures

Sub-diffraction resolution imaging has played a pivotal role in biological research by visualizing key, but previously unresolvable, sub-cellular structures. Unfortunately, applications of far-field sub-diffraction resolution are currently divided between fluorescent and coherent-diffraction regimes, and a multimodal sub-diffraction technique that bridges this gap has not yet been demonstrated. Here we report that structured illumination (SI) allows multimodal sub-diffraction imaging of both coherent quantitative-phase (QP) and fluorescence. Due to the conventional fluorescent applications of SI, we first demonstrated the principle of SI-enabled three-dimensional (3D) QP sub-diffraction imaging with calibration microspheres. Image analysis confirmed enhanced lateral and axial resolutions over diffraction-limited QP imaging, and established striking parallels between coherent SI and conventional optical diffraction tomography. We next introduce an optical system utilizing SI to achieve 3D sub-diffraction, multimodal QP/fluorescent visualization of A549 biological cells fluorescently tagged for F-actin. Our results suggest that SI has unique utility in studying biological phenomena with significant molecular, biophysical, and biochemical components., Comment: 15 pages of main text (including Figures), 13 pages of Supplementary Information (including Figures), 4 Figures in main text, 9 Figures in Supplementary Information