µOCT imaging using depth of focus extension by self-imaging wavefront division in a common-path fiber optic probe.

Optical coherence tomography (OCT) is an attractive medical modality due to its ability to acquire high-resolution, cross-sectional images inside the body using flexible, small-diameter, scanning fiber optic probes. Conventional, cross-sectional OCT imaging technologies have approximately 10-µm axial resolution and 30-µm lateral resolution, specifications that enable the visualization of microscopic architectural morphology. While this resolution is useful for many clinical applications, it is insufficient for resolving individual cells that characterize many diseases. To address this gap, a supercontinuum-laser-based, µm-resolution OCT (µOCT) system and a 500 µm-diameter, extended depth of focus single fiber optic probe for endoscopic and intravascular imaging were designed and fabricated. At the distal tip of the fiber optic probe, a cylindrical waveguide was used to divide the wavefront to provide multiple circular propagation modes. Once transmitted through a relatively high NA lens (NA >0.1), these modes were projected as multiple coaxial foci (~3 µm full width at half maximum (FWHM)) over a greatly extended focal depth range. The distal tip of the probe also contained a common-path reference reflectance to minimize polarization and dispersion imbalances between sample and reference arm light. Measurements showed that the probe provides a 20-fold depth of focus extension, maintaining a 3-5 µm lateral resolution (FWHM of PSF) and a 2 µm axial resolution over a depth range of approximately 1 mm. These results suggest that this new optical configuration will be useful for achieving high-resolution, cross-sectional OCT imaging in catheter/endoscope-based medical imaging devices.

Three-dimensional printing of tissue phantoms for biophotonic imaging.

We have investigated the potential of tissue phantoms fabricated with thermosoftening- and photopolymerization-based three-dimensional (3D) printers for use in evaluation of biophotonic imaging systems. The optical properties of printed polymer samples were measured and compared to biological tissues. Phantoms with subsurface channels as small as 0.2 mm in diameter were fabricated and imaged with microscopy, x-ray microtomography, and optical coherence tomography to characterize morphology. These phantoms were then implemented to evaluate the penetration depth of a hyperspectral reflectance imaging system used in conjunction with a near-infrared contrast agent. Results indicated that 3D printing may provide a suitable platform for performance testing in biophotonics, although subsurface imaging is critical to mitigate printer-to-printer variability in matrix homogeneity and feature microstructure.

Towards A Discretely Actuated Steerable Cannula for Diagnostic and Therapeutic Procedures.

We have designed, developed, and evaluated the performance of a multi-degree-of-freedom discretely actuated steerable cannula with shape memory alloy (SMA)actuators. This will enable us to deliver diagnostic as well as therapeutic devices to the target location through the hollow inner core of the cannula. We propose to use SMAs to generate bending forces due to its small size and high power density. We annealed the SMA wires through a customized training process in arc shape and mounted them at discrete locations on the outer surface of the cannula to enable joint motion. A pulse width modulation(PWM)-based control scheme was implemented to control all SMA actuators simultaneously to enable multiple joint motion using a single power supply. The proposed controller was validated through an experiment inside gelatin to mimic the motion of the cannula inside a medium which requires a significant amount of force to move the joints of the cannula. Trajectory planning using a suitable metric and trajectory execution were successfully implemented. To demonstrate the delivery of a diagnostic tool through our cannula, we demonstrate that we can pass an optical coherence tomography probe through the cannula and perform in situ micro-scale imaging.

A forward-imaging needle-type OCT probe for image guided stereotactic procedures.

A forward-imaging needle-type optical coherence tomography (OCT) probe with Doppler OCT (DOCT) capability has the potential to solve critical challenges in interventional procedures. A case in point is stereotactic neurosurgery where probes are advanced into the brain based on predetermined coordinates. Laceration of blood vessels in front of the advancing probe is an unavoidable complication with current methods. Moreover, cerebrospinal fluid (CSF) leakage during surgery can shift the brain rendering the predetermined coordinates unreliable. In order to address these challenges, we developed a forward-imaging OCT probe (740 µm O.D.) using a gradient-index (GRIN) rod lens that can provide real-time imaging feedback for avoiding at-risk vessels (8 frames/s with 1024 A-scans per frame for OCT/DOCT dual imaging) and guiding the instrument to specific targets with 12 µm axial resolution (100 frames/s with 160 A-scans per frame for OCT imaging only). The high signal-to-background characteristic of DOCT provides exceptional sensitivity in detecting and quantifying the blood flow within the sheep brain parenchyma in real time. The OCT/DOCT dual imaging also demonstrated its capability to differentiate the vessel type (artery/vein) on rat's femoral vessels. We also demonstrated in ex vivo human brain that the location of the tip of the OCT probe can be inferred from micro-anatomical landmarks in OCT images. These findings demonstrate the suitability of OCT guidance during stereotactic procedures in the brain and its potential for reducing the risk of cerebral hemorrhage.

Optical coherence tomography-guided laser marking with tethered capsule endomicroscopy in unsedated patients.

Tethered capsule endomicroscopy (TCE) is an emerging screening technology that comprehensively obtains microstructural OCT images of the gastrointestinal (GI) tract in unsedated patients. To advance clinical adoption of this imaging technique, it will be important to validate TCE images with co-localized histology, the current diagnostic gold standard. One method for co-localizing OCT images with histology is image-targeted laser marking, which has previously been implemented using a driveshaft-based, balloon OCT catheter, deployed during endoscopy. In this paper, we present a TCE device that scans and targets the imaging beam using a low-cost stepper motor that is integrated inside the capsule. In combination with a 4-laser-diode, high power 1430/1450 nm marking laser system (800 mW on the sample and 1s pulse duration), this technology generated clearly visible marks, with a spatial targeting accuracy of better than 0.5 mm. A laser safety study was done on swine esophagus ex vivo, showing that these exposure parameters did not alter the submucosa, with a large, 4-5x safety margin. The technology was demonstrated in living human subjects and shown to be effective for co-localizing OCT TCE images to biopsies obtained during subsequent endoscopy.

Quantitative single-mode fiber based PS-OCT with single input polarization state using Mueller matrix.

We present a simple but effective method to quantitatively measure the birefringence of tissue by an all single-mode fiber (SMF) based polarization-sensitive optical coherence tomography (PS-OCT) with single input polarization state. We theoretically verify that our SMF based PS-OCT system can quantify the phase retardance and optic axis orientation after a simple calibration process using a quarter wave plate (QWP). Based on the proposed method, the quantification of the phase retardance and optic axis orientation of a Berek polarization compensator and biological tissues were demonstrated.

Real-time epidural anesthesia guidance using optical coherence tomography needle probe.

Epidural anesthesia is one of the most widely used anesthesia methods. Due to lack of visual feedback to guide needle navigation, failure rate of epidural anesthesia is up to 20%, and the complication rate of peripheral nerve block approaches 10%, with the potential of permanent nerve damage. To address these difficulties, needle insertion under ultrasound guidance and fluoroscopy has been introduced. However, they do not provide adequate resolution and contrast to distinguish the tissue layers that the needle travels through or to specifically identify the epidural space. To improve the accuracy of epidural space identification, we developed a small hand-held optical coherence tomography (OCT) forward-imaging needle device for real-time epidural anesthesia surgery guidance and demonstrated its feasibility through ex vivo and in vivo animal experiments. With tissue structures visualized and differentiated at the needle tip, OCT needle imaging device will enhance clinical outcomes with regards to complication rates, induced pain, and procedure failure when compared to standard practice. Furthermore, this technology could be used in combination with ultrasound/fluoroscopy to enhance outcomes.

In Vivo Voltage-Sensitive Dye Imaging of Subcortical Brain Function.

The whisker system of rodents is an excellent model to study peripherally evoked neural activity in the brain. Discrete neural modules represent each whisker in the somatosensory cortex ("barrels"), thalamus ("barreloids"), and brain stem ("barrelettes"). Stimulation of a single whisker evokes neural activity sequentially in its corresponding barrelette, barreloid, and barrel. Conventional optical imaging of functional activation in the brain is limited to surface structures such as the cerebral cortex. To access subcortical structures and image sensory-evoked neural activity, we designed a needle-based optical system using gradient-index (GRIN) rod lens. We performed voltage-sensitive dye imaging (VSDi) with GRIN rod lens to visualize neural activity evoked in the thalamic barreloids by deflection of whiskers in vivo. We stimulated several whiskers together to determine the sensitivity of our approach in differentiating between different barreloid responses. We also carried out stimulation of different whiskers at different times. Finally, we used muscimol in the barrel cortex to silence the corticothalamic inputs while imaging in the thalamus. Our results show that it is possible to obtain functional maps of the sensory periphery in deep brain structures such as the thalamic barreloids. Our approach can be broadly applicable to functional imaging of other core brain structures.

Real-time monitoring of hemodynamic changes in tumor vessels during photoimmunotherapy using optical coherence tomography.

Photoimmunotherapy (PIT) is a cell-specific cancer therapy based on an armed antibody conjugate that induces rapid and highly selective cancer cell necrosis after exposure to near-infrared (NIR) light. The PIT treatment also induces the superenhanced permeability and retention effect, which allows high concentrations of nanoparticles to accumulate in the tumor bed. In our pilot studies, optical coherence tomography (OCT) reveals dramatic hemodynamic changes during PIT. We developed and applied speckle variance analysis, Doppler flow measurement, bulk motion removal, and automatic region of interest selection to quantify vessel diameter and blood velocity within tumors in vivo. OCT imaging reveals that blood velocity in peripheral tumor vessels quickly drops below the detection limit while the vessel lumen remains open (4 vessels from 3 animals). On the other hand, control tumor vessels (receive NIR illumination but no PIT drug) do not show the sustained blood velocity drop (5 vessels from 3 animals). Ultraslow blood velocity could result in a long drug circulation time in tumor. Increase of the blood pool volume within the central tumor (shown in histology) may be the leading cause of the periphery blood velocity drop and could also increase the drug pool volume in tumor vessels.

Concurrent multiscale imaging with magnetic resonance imaging and optical coherence tomography.

We develop a novel platform based on a tele-operated robot to perform high-resolution optical coherence tomography (OCT) imaging under continuous large field-of-view magnetic resonance imaging (MRI) guidance. Intra-operative MRI (iMRI) is a promising guidance tool for high-precision surgery, but it may not have sufficient resolution or contrast to visualize certain small targets. To address these limitations, we develop an MRI-compatible OCT needle probe, which is capable of providing microscale tissue architecture in conjunction with macroscale MRI tissue morphology in real time. Coregistered MRI/OCT images on ex vivo chicken breast and human brain tissues demonstrate that the complementary imaging scales and contrast mechanisms have great potential to improve the efficiency and the accuracy of iMRI procedure.

Coherence-gated Doppler: a fiber sensor for precise localization of blood flow.

Miniature optical sensors that can detect blood vessels in front of advancing instruments will significantly benefit many interventional procedures. Towards this end, we developed a thin and flexible coherence-gated Doppler (CGD) fiber probe (O.D. = 0.125 mm) that can be integrated with minimally-invasive tools to provide real-time audio feedback of blood flow at precise locations in front of the probe. Coherence-gated Doppler (CGD) is a hybrid technology with features of laser Doppler flowmetry (LDF) and Doppler optical coherence tomography (DOCT). Because of its confocal optical design and coherence-gating capabilities, CGD provides higher spatial resolution than LDF. And compared to DOCT imaging systems, CGD is simpler and less costly to produce. In vivo studies of rat femoral vessels using CGD demonstrate its ability to distinguish between artery, vein and bulk movement of the surrounding soft tissue. Finally, by placing the CGD probe inside a 30-gauge needle and advancing it into the brain of an anesthetized sheep, we demonstrate that it is capable of detecting vessels in front of advancing probes during simulated stereotactic neurosurgical procedures. Using simultaneous ultrasound (US) monitoring from the surface of the brain we show that CGD can detect at-risk blood vessels up to 3 mm in front of the advancing probe. The improved spatial resolution afforded by coherence gating combined with the simplicity, minute size and robustness of the CGD probe suggest it may benefit many minimally invasive procedures and enable it to be embedded into a variety of surgical instruments.

Characterizing the point spread function of retinal OCT devices with a model eye-based phantom.

We have designed, fabricated, and tested a nanoparticle-embedded phantom (NEP) incorporated into a model eye in order to characterize the point spread function (PSF) of retinal optical coherence tomography (OCT) devices in three dimensions under realistic imaging conditions. The NEP comprises a sparse distribution of highly backscattering silica-gold nanoshells embedded in a transparent UV-curing epoxy. The commercially-available model eye replicates the key optical structures and focusing power of the human eye. We imaged the model eye-NEP combination with a research-grade spectral domain OCT system designed for in vivo retinal imaging and quantified the lateral and axial PSF dimensions across the field of view in the OCT images. We also imaged the model eye-NEP in a clinical OCT system. Subtle features in the PSF and its dimensions were consistent with independent measurements of lateral and axial resolution. This model eye-based phantom can provide retinal OCT device developers and users a means to rapidly, objectively, and consistently assess the PSF, a fundamental imaging performance metric.

Review of advanced imaging techniques.

Pathology informatics encompasses digital imaging and related applications. Several specialized microscopy techniques have emerged which permit the acquisition of digital images ("optical biopsies") at high resolution. Coupled with fiber-optic and micro-optic components, some of these imaging techniques (e.g., optical coherence tomography) are now integrated with a wide range of imaging devices such as endoscopes, laparoscopes, catheters, and needles that enable imaging inside the body. These advanced imaging modalities have exciting diagnostic potential and introduce new opportunities in pathology. Therefore, it is important that pathology informaticists understand these advanced imaging techniques and the impact they have on pathology. This paper reviews several recently developed microscopic techniques, including diffraction-limited methods (e.g., confocal microscopy, 2-photon microscopy, 4Pi microscopy, and spatially modulated illumination microscopy) and subdiffraction techniques (e.g., photoactivated localization microscopy, stochastic optical reconstruction microscopy, and stimulated emission depletion microscopy). This article serves as a primer for pathology informaticists, highlighting the fundamentals and applications of advanced optical imaging techniques.