Bioimaging and Biomedical Optics Commons^™

A Comparison Of Optical Measurement Methods For The Growth Of S. Cerevisiae, Jackson Black

Chemical Engineering Undergraduate Honors Theses

Genetic engineering of living organisms provides the opportunity to express and harvest different proteins from cell surfaces. Yeast (S. cerevisiae) is one such organism and is capable of being grown on an industrial scale. Cellular concentration is an important parameter to monitor while fermentation processes are underway, in order to control the environment inside the growth medium and maximize yields. Spectrophotometry is a conventional method for measuring concentration, but is limited by a narrow absorbance range, and the need for on-site periodic sampling. A continuous method of measurement, as provided by Bug Labs BE2100 non-invasive biomass monitor, would …

Synthesis, Radiolabeling And Evaluation Of A Suite Of Tracers With 44Sc For Detecting Extracellular Dna, Zhiyao Li

McKelvey School of Engineering Graduate Student Theses & Dissertations

Neutrophil extracellular traps involve the rapid translocation of DNA to the outside of the cell under certain stimuli. This structure forms a fibrous network that is able to limit the spread of pathogens and to kill microorganisms. It has also been shown to be present in various pathological processes such as inflammation, autoimmune diseases, and cancer metastasis. Currently, the formation process of NETs in vivo is being extensively studied. However noninvasive detection and quantitation has yet to be achieved. A class of PET tracers are described here that consists of a DNA dye as the backbone that is labeled with …

Light Sheet Microscopy Incubation Design And Control Scheme, Venkatanathan Kidambi

Honors Scholar Theses

The focus of the following paper is the development of a light sheet or selective plane illumination microscopy (LSM / SPIM) platform to support long duration three dimensional imaging of various cells and tissues. This paper describes the complete design and fabrication process behind the prototype, including a definition of light sheet microscopy and its differences from confocal microscopy, as well as existing open source selective plane microscopy literature and designs. The complete engineering design process as applied to the microscope design is described in detail. Specifically, this paper will focus on the development of an incubation chamber for light …

Automated Segmentation Of Autofluorescence Lifetime Images Using A Convolutional Neural Network, Nicole Hooten

Biomedical Engineering Undergraduate Honors Theses

Chronic wounds affect nearly 1 out of every 50 people in the United States, decreasing quality of life and putting people with potential comorbidities at high risk for obtaining an infection. Wound healing progress in clinical settings is measured by tracking wound size and there are currently no non-invasive, quantitative measurement techniques. Previous studies have proposed nicotinamide adenine dinucleotide (NADH) autofluorescence lifetime (FLIM) imaging of the wound edge as a method to quantify the wound healing process by connecting the role of NADH in cellular metabolism to wound healing stage. However, evaluation of FLIM images is heavily subjective, as a …

Portable Diffuse Reflectance Spectroscopy For Non-Invasive And Quantitative Assessment Of The Parathyroid Glands Viability During Surgery, Mark Romine, Linh Luong, Alex Moazzen, Katie Cho, Paul Lee

Symposium of Student Scholars

Portable Diffuse Reflectance Spectroscopy for Non-invasive and Quantitative Assessment of the Parathyroid Glands Viability During Surgery

Mark Romine, Linh Luong, Alex Moazzen, Katie Cho and Paul Lee

The parathyroid glands (PTGs) are responsible for the regulation of calcium levels in the blood by secreting a parathyroid hormone. This parathyroid hormone then regulates the body’s absorption, storage, and secretion of calcium, which can directly affect the way muscles and nerves operate. PTGs are often at risk of damage, or accidental removal during thyroid surgeries, because it is challenging to identify PTGs and to determine their viability. Current methods of visual inspections …

Wireless, Handheld Diffuse Reflectance Spectroscopy To Quantify Tissue Microvascular Hemodynamics, Linh Luong, Alex Moazzen, Mark Romine, Katie Cho, Paul Lee

Symposium of Student Scholars

Diffuse Reflectance Spectroscopy (DRS) is a non-invasive optical method to characterize tissue optical properties for disease diagnosis and health monitoring. Two optical fibers are often used in a DRS system: one to deliver light to the tissue and the other to gather diffuse reflectance spectra, which provide quantitative details about the structure and composition of the tissue. The conventional DRS system, however, is expensive, bulky, and composed of fragile optical fibers and multiple electrical connections. Here we propose to build a wireless, handheld, and fiber-less diffuse optical spectroscopy system. Unfortunately, the diffusion approximation utilized for data analysis of the conventional …

Enhancement Of Deep Learning Protein Structure Prediction, Ruoming Shen

Modeling, Simulation and Visualization Student Capstone Conference

Protein modeling is a rapidly expanding field with valuable applications in the pharmaceutical industry. Accurate protein structure prediction facilitates drug design, as extensive knowledge about the atomic structure of a given protein enables scientists to target that protein in the human body. However, protein structure identification in certain types of protein images remains challenging, with medium resolution cryogenic electron microscopy (cryo-EM) protein density maps particularly difficult to analyze. Recent advancements in computational methods, namely deep learning, have improved protein modeling. To maximize its accuracy, a deep learning model requires copious amounts of up-to-date training data.

This project explores DeepSSETracer, a …

Image Analysis Methodology For Evaluating Coating Uniformity Of Drug-Coated Angioplasty Balloons, Allison Dobson Zieschang

LSU Master's Theses

Percutaneous transluminal balloon angioplasty (PTA) is a surgical procedure to treat peripheral artery disease, but restenosis causes suboptimal outcomes in many patients. Drug-coated angioplasty balloons (DCBs) were developed to reduce the risk of restenosis by delivering anti-proliferative drugs to the lesion. The efficacy of DCBs depends on factors like drug loading and drug release rate, both of which are directly impacted by the coating uniformity. This thesis evaluates the uniformity of DBCs using fluorescence microscopy, scanning electron microscopy (SEM), qualitative analysis, and an image processing algorithm. Uniformity was quantified by three measurements: histogram standard deviation, histogram percentage of coverage, and …

Computational Design Of Fiber-Optic Probes For Biosensing, Suwarna Karna

Electrical Engineering Theses

This thesis presents a study on the optical characteristics of hollow-core photonic crystal fibers (HC-PCFs) with a band gap cladding structure and their applications in optical fiber sensing. This 800B HC-PCF exhibited excellent optical properties and has a flexible structure, which makes them suitable for a wide range of industrial applications. Finite element simulations and structural optimization designs were conducted using the surface plasmon resonance (SPR) technique to determine the optimal performance parameters of the 800B HC-PCF. The fiber was further modified using the SPR technique to improve its practical detection capabilities. The performance of the modified fiber was observed …

A Device For Measuring Acoustic Output Intensity Of Transcranial Doppler Ultrasound Transducers For Comparison With Fda Regulations, Sarah Altman, Gregory Bashford, Benjamin Hage

Honors Program: Senior Projects (Public)

Transcranial Doppler Ultrasonography (TCD) is a non-invasive methodology which can evaluate cerebral blood flow velocity in real time. Single-element focused circular transducers placed on the scalp produce ultrasound waves capable of penetrating the skull with minimal aberration, enabling measurement of the Doppler shift. As such, TCD can measure blood flow velocity in the internal carotid and vertebral arteries, as well as the arteries of the Circle of Willis: the Anterior Cerebral (ACA), Posterior Cerebral (PCA), and Middle Cerebral Arteries (MCA). Intracranial aneurysm and ischemic stroke are serious conditions in which ballooning or occlusion of cerebral vessels cause insufficient perfusion, leading …

Differentiating Axonal From Demyelinating Neuropathies Using Multiparametric Quantitative Mri Of Peripheral Nerves, Jacob D. Baraz, Stephanie Xuan, Sadaf Saba, Xue Yang, Ryan Castoro, Yang Xuan, Alison Roth, Richard D. Dortch, Jun Li, Yongsheng Chen

Medical Student Research Symposium

Objectives: To develop a multiparametric quantitative MRI (qMRI) method to track pathological changes in the peripheral neuropathies.

Background: Irrespective of the causes or types of polyneuropathies, peripheral nerves are mainly afflicted by two kinds of pathologies – axonal loss and demyelination. It is critical to differentiate between the two as treatments are different for the two conditions. While nerve conduction studies (NCS) have been used to differentiate the two pathologies in the distal nerves, there are no tools to probe the pathologies in the proximal peripheral nerves. This is particularly needed when distal nerves become non-responsive in NCS.

Methods: We …

Efficient Scopeformer: Towards Scalable And Rich Feature Extraction For Intracranial Hemorrhage Detection Using Hybrid Convolution And Vision Transformer Networks, Yassine Barhoumi

Theses and Dissertations

The field of medical imaging has seen significant advancements through the use of artificial intelligence (AI) techniques. The success of deep learning models in this area has led to the need for further research. This study aims to explore the use of various deep learning algorithms and emerging modeling techniques to improve training paradigms in medical imaging. Convolutional neural networks (CNNs) are the go-to architecture for computer vision problems, but they have limitations in mapping long-term dependencies within images. To address these limitations, the study explores the use of techniques such as global average pooling and self-attention mechanisms. Additionally, the …

Deutj: An Imagej Plugin For Improved Automatic Masking And Segmentation Of Images From Confocal Microscopy, Sunny Cui

Independent Student Projects and Publications

Due to advances in microscopic imaging, there are a plethora of biological molecules that can now be tagged and subsequentially imaged from almost any cell, organism, or tissue. However, the ability of software to analyze these images remains to be a challenge. ImageJ is open source software that allows for the processing of these images, but faces challenges when dealing with images that show weak contrast between objects of interest and background. DeutJ is an ImageJ plugin that color corrects and brightness corrects gradients that inhibit segmentation in confocal microscopy images. It can analyze hundreds of images from a given …

Session 12: Active Learning To Minimize The Possible Risk From Future Epidemics, Kc Santosh

SDSU Data Science Symposium

In medical imaging informatics, for any future epidemics (e.g., Covid-19), deep learning (DL) models are of no use as they require a large dataset as they take months and even years to collect enough data (with annotations). In such a context, active learning (or human/expert-in-the-loop) is the must, where a machine can learn from the first day with minimum possible labeled data. In unsupervised learning, we propose to build pre-trained DL models that iteratively learn independently over time, where human/expert intervenes only when it makes mistakes and for only a limited data. In our work, deep features are used to …

Brain Activity Associated With Taste Stimulation: A Mechanism For Neuroplastic Change?, Angela M. Dietsch, Ross M. Westemeyer, Douglas H. Schultz

Department of Special Education and Communication Disorders: Faculty Publications

Purpose: Neuroplasticity may be enhanced by increasing brain activation and bloodflow in neural regions relevant to the target behavior.We administered precisely formulated and dosed taste stimuli to determine whether the associated brain activity patterns included areas that underlie swallowing control.

Methods: Five taste stimuli (unflavored, sour, sweet-sour, lemon, and orange suspensions) were administered in timing-regulated and temperature-controlled 3 mL doses via a customized pump/tubing system to 21 healthy adults during functional magnetic resonance imaging (fMRI). Whole-brain analyses of fMRI data assessed main effects of taste stimulation as well as differential effects of taste profile.

Results: Differences in …

Estimation Of Free Water-Corrected Microscopic Fractional Anisotropy., Nico J J Arezza, Tales Santini, Mohammad Omer, Corey A Baron

Medical Biophysics Publications

Water diffusion anisotropy MRI is sensitive to microstructural changes in the brain that are hallmarks of various neurological conditions. However, conventional metrics like fractional anisotropy are confounded by neuron fiber orientation dispersion, and the relatively low resolution of diffusion-weighted MRI gives rise to significant free water partial volume effects in many brain regions that are adjacent to cerebrospinal fluid. Microscopic fractional anisotropy is a recent metric that can report water diffusion anisotropy independent of neuron fiber orientation dispersion but is still susceptible to free water contamination. In this paper, we present a free water elimination (FWE) technique to estimate microscopic …

Dataset For Manuscript: Comparing Performance Of Spectral Image Analysis Approaches For Detection Of Cellular Signals In Time-Lapse Hyperspectral Imaging Fluorescence Excitation-Scanning Microscopy, Silas J. Leavesley

BioImaging and BioSystems Research

The dataset contains raw and processed hyperspectral timelapse image data that are described in the manuscript:

Parker, M., Annamdevula, N. S., Pleshinger, D., Ijaz, Z., Jalkh, J., Penn, R., Deshpande, D., Rich, T. C. & Leavesley, S. J. Comparing Performance of Spectral Image Analysis Approaches for Detection of Cellular Signals in Time-Lapse Hyperspectral Imaging Fluorescence Excitation-Scanning Microscopy. Bioengineering 10, 642 (2023).

Advanced Modeling And Innovative Transcranial Magnetic Stimulation Coil Designs For The Treatment Of Neurological Disorders, Mohannad Tashli

Theses and Dissertations

Transcranial magnetic stimulation (TMS) is a safe, effective and non-invasive treatment for several mental and psychiatric disorders. TMS is an FDA approved treatment and is commonly applied to patients who do not respond to medications for the treatment of clinical depression, smoking cessation, obsessive-compulsive disorder and migraine. Recently, there has been an increase in the development of electromagnetic neuromodulation techniques targeted at enhancing the effectiveness of TMS devices for the treatment of mental diseases. In TMS stimulation, focality is an important factor which determines the specificity of the pulses induced in different brain tissues. The electromagnetic pulses must be confined …

Multiscale And Multimodality Optical Imaging Of Brain Hemodynamics And Function, Mehrana Mohtasebi

Theses and Dissertations--Biomedical Engineering

Frequent measurement of cerebral hemodynamics is crucial for continuous assessment of brain function and health. Low-frequency oscillations (LFOs; < 0.1 Hz) have shown promise as an indicator of altered neurologic activity in the abnormal brain. Moreover, quantification of temporal correlations in LFOs across distinct brain regions enables mapping of regional neurovascular disorders. Portable diffuse optical techniques such as near-infrared spectroscopy (NIRS) and diffuse correlation spectroscopy (DCS) enable noninvasive and continuous measurements of cerebral oxygenation and cerebral blood flow (CBF), respectively. However, most NIRS/DCS systems use limited numbers of discrete light sources and detectors and thus lack combinations of high temporal/spatial resolution and large field-of-view (FOV) for brain imaging. Recent development of an innovative noncontact speckle contrast diffuse correlation tomography (scDCT) in our lab provides a low-cost and portable tool for high-density imaging of CBF distributions over large FOVs. The scDCT uses a galvo-mirror to remotely deliver coherent point near-infrared light to multiple source positions and a CMOS camera as a high-density 2-D detector array to quantify spatial fluctuations of diffuse laser speckles, resulting from movement of red blood cells. Boundary CBF data were input to a unique finite-element-method (FEM)-based reconstruction algorithm for 3-D imaging of CBF distributions. However, scDCT was hindered for extracting LFOs due to its low sampling rate. In this study, scDCT was optimized to achieve high sampling rates and fast image reconstructions by use of moving window algorithms with parallel computations (Project 1). Power spectral density (PSD) analysis was performed to investigate altered LFOs during transient cerebral ischemia in neonatal piglets. Transient cerebral ischemia caused reductions in both CBF and PSD, supporting inclusion of scDCT in the growing field of LFO analysis. One remaining limitation was the long computational time required for 3-D reconstruction, which prohibited real-time applications. To overcome this limitation, a depth-sensitive diffuse speckle contrast topography (DSCT) method was developed for analyzing scDCT data to achieve fast and high-density 2-D mapping of CBF distribution without the need of complex and time-consuming 3-D reconstructions (Project 2). New head-simulating phantoms with known optical/geometric properties were designed and fabricated to evaluate the spatial resolution and depth sensitivity of DSCT method. In-vivo experiments verified the capability of DSCT in tracking dynamic changes in CBF at different depths during CO₂ inhalation and transient cerebral ischemia in rats. In addition to preclinical studies, a low-cost, wearable, fluorescence eye loupe (FLoupe) device was invented for intraoperative visualization of brain tumor margin in the clinic (Project 3). Consistent results were observed against an expensive, large, standard fluorescence operative microscope for the guidance of tumor resection. Overall, through 3 projects this dissertation demonstrated a variety of noninvasive noncontact optical technologies (scDCT, DSCT, and FLoupe) that enabled multiscale (rodents, piglets, and humans) and …

A Wearable Fiber-Free Optical Sensor For Continuous Measurements Of Cerebral Blood Flow And Oxygenation, Xuhui Liu

Theses and Dissertations--Biomedical Engineering

Wearable technologies for continuous monitoring of cerebral hemodynamics in freely behaving subjects not only advance our understanding of cognitive processing and adaptive behavior, but also provide vital information for diagnosis and therapeutic assessment of cerebral diseases associated with hypoxia/ischemia. Wearable near-infrared diffuse optical techniques have been used at the bedside to probe deep cerebral hemodynamics, including near-infrared spectroscopy (NIRS) for cerebral oxygenation measurement and diffuse correlation spectroscopy (DCS) for cerebral blood flow (CBF) measurement, respectively. However, most systems are relatively large and expensive, and use rigid fiber-optic probes that significantly constrain subject’s movement. A novel, wearable, fiber-free diffuse speckle contrast …

Using Paired-Agent Imaging To Track Changes In Head And Neck Cancers After Undergoing Photodynamic Therapy Treatment, Reeham M. Choudhury

Dartmouth College Master’s Theses

Head and neck cancers affect thousands of people across the world, and photodynamic therapy (PDT) has been shown to have great potential to treat said cancers in a noninvasive manner. However, imaging head and neck cancers has been difficult, and molecular changes caused by PDT are not well-understood. Therefore, we propose the use of paired-agent imaging (PAI) to track changes in these cancers after after PDT treatment. For these studies, we primarily used benzoporphyrin derivative monoacid (BPD) for our photosensitizer, which is activated by a 690 nm laser. We first looked at changes in EGFR expression in vitro, and …

Evaluating Human Eye Features For Objective Measure Of Working Memory Capacity, Yasasi Abeysinghe, Enkelejda Kasneci (Ed.), Frederick Shic (Ed.), Mohamed Khamis (Ed.)

Computer Science Faculty Publications

Eye tracking measures can provide means to understand the underlying development of human working memory. In this study, we propose to develop machine learning algorithms to find an objective relationship between human eye movements via oculomotor plant and their working memory capacity, which determines subjective cognitive load. Here we evaluate oculomotor plant features extracted from saccadic eye movements, traditional positional gaze metrics, and advanced eye metrics such as ambient/focal coefficient , gaze transition entropy, low/high index of pupillary activity (LHIPA), and real-time index of pupillary activity (RIPA). This paper outlines the proposed approach of evaluating eye movements for obtaining an …

Ultrasensitive Tapered Optical Fiber Refractive Index, Erem Ujah, Meimei Lai, Gymama Slaughter

Electrical & Computer Engineering Faculty Publications

Refractive index (RI) sensors are of great interest for label-free optical biosensing. A tapered optical fiber (TOF) RI sensor with micron-sized waist diameters can dramatically enhance sensor sensitivity by reducing the mode volume over a long distance. Here, a simple and fast method is used to fabricate highly sensitive refractive index sensors based on localized surface plasmon resonance (LSPR). Two TOFs (l = 5 mm) with waist diameters of 5 µm and 12 µm demonstrated sensitivity enhancement at λ = 1559 nm for glucose sensing (5-45 wt%) at room temperature. The optical power transmission decreased with increasing glucose concentration due …

Verification Of Proton Range Predictions In Proton Treatment Planning Using X-Ray Ct Or Proton Ct Imaging, Nicholas S. Yee

Graduate Research Theses & Dissertations

The objective of this paper is to compare the range errors for two treatment plans; one using x-ray CT and the other using proton CT for calculating dose distributions in an animal tissue phantom. We shot 150 MeV proton beam through a meat phantom to a film stack to measure the exponential drop of dose at the Bragg peak until zero dose remained. Plotting the depth dose curves and performing gamma analyses are two ways to quantify the proton range errors when comparing them to depth dose curves from film measurements in the phantom. The depth dose curves give us …

Patient-Specific Analysis Of Aortic Hemodynamics And Wall Shear Stresses In Patients Undergoing Pediatric Bariatric Surgery Using 2d Phase-Contrast Mri, Oluwaferanmi S. Akande, Joao Soares, Nathan Hargan, Dr. Uyen Truong, Haeung Kang, David Lanning

Undergraduate Research Posters

Patient-Specific Analysis of Aortic Hemodynamics and Wall Shear Stresses in Patients Undergoing Pediatric Bariatric Surgery using 2D Phase-Contrast MRI

Feranmi Akande¹ Nathan Hargan² Haesung Kang³ David Lanning³ Uyen Truong⁴ João S. Soares¹

¹ Department of Biomedical Engineering, College of Engineering, Virginia Commonwealth University

² Department of Mechanical & Nuclear Engineering, College of Engineering, Virginia Commonwealth University

³ Department of Cardiology, Children’s Hospital of Richmond, Virginia Commonwealth University

Introduction: Severe obesity among today’s youth is a rising epidemic within the United States. Emerging research has linked childhood obesity with shortened life expectancy and early …

Soft X-Ray And Susceptibility Based Magnetic Resonance Imaging To Map Iron Distribution In Apples: Initial Results To Model Iron Storage In Water-Deficient Or Dehydrated Biological Tissue, Subhendra Sarkar, Eric Lobel, Evans Lespinasse, Zoya Vinokur, Analia Basilicata, Sonia Orellana, Maria Orellana, Aaliyah Salmon, Joanna Syska, Aravis Mcbroom, Jian Wang, Anam Riaz, Jody-Ann Douglas

Publications and Research

Radiology departments have contributed significantly to greenhouse gases including release of toxic imaging contrast media to environment. We feel Radiology also has several spectroscopy and imaging tools that may apply to monitor and support cleaner environmental goals. The current manuscript is one of the firsts to prompt Radiology to move in that direction by non-invasive imaging of bio metals that are less abundant in biological tissues but play key roles as co-factors in tissue structure and function. Conventional analytical tools are mostly invasive and cannot characterize the native oxidation states of bio metals. We chose carbohydrate matrix of metal-rich fruits …

Measurement Of Fluid Movement Throughout The Brain Via Multiple Magnetic Resonance Imaging Techniques During High-Definition Transcranial Direct Current Stimulation, Jack W. Beaty Mr

Dissertations and Theses

Transcranial Direct Current Stimulation (tDCS) is a non-invasive method of neuromodulation with applications in neuroscience, clinical care, and biomedical engineering. tDCS has been well established as a safe method of applying low amplitude current between two or more electrodes to alter excitation thresholds and neuroplasticity. Recently, in-vitro and clinical studies have suggested that DC stimulation can induce a transient, polarity-specific, effect of increased water exchange across the blood-brain barrier (BBB). The electroosmotic effect, the proposed phenomenon driving water exchange, is a biophysical response of charged ions moving across an oppositely-charged surface, i.e., the tight junction, when subjected to an electric …

Advancing Fluorescent Contrast Agent Recovery Methods For Surgical Guidance Applications, Brook Kennedy Byrd

Dartmouth College Ph.D Dissertations

Fluorescence-guided surgery (FGS) utilizes fluorescent contrast agents and specialized optical instruments to assist surgeons in intraoperatively identifying tissue-specific characteristics, such as perfusion, malignancy, and molecular function. In doing so, FGS represents a powerful surgical navigation tool for solving clinical challenges not easily addressed by other conventional imaging methods. With growing translational efforts, major hurdles within the FGS field include: insufficient tools for understanding contrast agent uptake behaviors, the inability to image tissue beyond a couple millimeters, and lastly, performance limitations of currently-approved contrast agents in accurately and rapidly labeling disease. The developments presented within this thesis aim to address such …

Small-Separation Speckle Contrast Optical Spectroscopy For Intraoperative Assessment Of Parathyroid Glands Viability During Thyroid Surgery, Connor Berger

Symposium of Student Scholars

The parathyroid glands (PTGs) are often damaged during thyroid surgeries due to a lack of methods identifying PTGs and assessing their viability. Damage to PTGs can cause hypocalcemia, a deficiency of calcium in the body. This complication can lead to detrimental consequences with economic burden. The surgeon’s current method of viability assessment is qualitative and subjective. Our technical solution is to employ an optical technique called speckle contrast optical spectroscopy (SCOS) that noninvasively quantifies the blood flow index (Db) of biological tissues at deep tissue levels (>1cm). The goal of this project is to verify SCOS at small source-detector-separation …

Six-Dimensional Single-Molecule Imaging With Isotropic Resolution Using A Multi-View Reflector Microscope, Oumeng Zhang, Zijian Guo, Yuanyuan He, Tingting Wu, Michael D. Vahey, Matthew D. Lew

Electrical & Systems Engineering Publications and Presentations

Imaging of both the positions and orientations of single fluorophores, termed single-molecule orientation-localization microscopy, is a powerful tool for the study of biochemical processes. However, the limited photon budget associated with single-molecule fluorescence makes high-dimensional imaging with isotropic, nanoscale spatial resolution a formidable challenge. Here we realize a radially and azimuthally polarized multi-view reflector (raMVR) microscope for the imaging of the three-dimensional (3D) positions and 3D orientations of single molecules, with precisions of 10.9 nm and 2.0° over a 1.5-μm depth range. The raMVR microscope achieves 6D super-resolution imaging of Nile red molecules transiently bound to lipid-coated spheres, accurately resolving …