Jeff Fingler

Mobility and transverse flow visualization using phase variance contrast with spectral domain optical coherence tomography

Phase variance-based motion contrast is demonstrated using two phase analysis methods in a spectral domain optical coherence tomography system. Mobility contrast is demonstrated for an intensity matched Intralipid solution placed without flow within agarose wells. Vasculature oriented transversely to the imaging direction has been imaged for 3-4 dpf in vivo zebrafish using the phase variance contrast methods. 2D phase variance contrast images are demonstrated with imaging times only 25% higher than a Doppler flow image with comparable statistics. En face images created by integrating depth regions of 3D zebrafish intensity and phase variance contrast data demonstrate vasculature consistent with expected images.

In vivo volumetric imaging of human retinal circulation with phase-variance optical coherence tomography.

We present in vivo volumetric images of human retinal micro-circulation using Fourier-domain optical coherence tomography (Fd-OCT) with the phase-variance based motion contrast method. Currently fundus fluorescein angiography (FA) is the standard technique in clinical settings for visualizing blood circulation of the retina. High contrast imaging of retinal vasculature is achieved by injection of a fluorescein dye into the systemic circulation. We previously reported phase-variance optical coherence tomography (pvOCT) as an alternative and non-invasive technique to image human retinal capillaries. In contrast to FA, pvOCT allows not only noninvasive visualization of a two-dimensional retinal perfusion map but also volumetric morphology of retinal microvasculature with high sensitivity. In this paper we report high-speed acquisition at 125 kHz A-scans with pvOCT to reduce motion artifacts and increase the scanning area when compared with previous reports. Two scanning schemes with different sampling densities and scanning areas are evaluated to find optimal parameters for high acquisition speed in vivo imaging. In order to evaluate this technique, we compare pvOCT capillary imaging at 3x3 mm2 and 1.5x1.5 mm2 with fundus FA for a normal human subject. Additionally, a volumetric view of retinal capillaries and a stitched image acquired with ten 3x3 mm2 pvOCT sub-volumes are presented. Visualization of retinal vasculature with pvOCT has potential for diagnosis of retinal vascular diseases.

Volumetric microvascular imaging of human retina using optical coherence tomography with a novel motion contrast technique

Phase variance-based motion contrast imaging is demonstrated using a spectral domain optical coherence tomography system for the in vivo human retina. This contrast technique spatially identifies locations of motion within the retina primarily associated with vasculature. Histogram-based noise analysis of the motion contrast images was used to reduce the motion noise created by transverse eye motion. En face summation images created from the 3D motion contrast data are presented with segmentation of selected retinal layers to provide non-invasive vascular visualization comparable to currently used invasive angiographic imaging. This motion contrast technique has demonstrated the ability to visualize resolution-limited vasculature independent of vessel orientation and flow velocity.

Optical imaging of the chorioretinal vasculature in the living human eye

Detailed visualization of microvascular changes in the human retina is clinically limited by the capabilities of angiography imaging, a 2D fundus photograph that requires an intravenous injection of fluorescent dye. Whereas current angiography methods enable visualization of some retinal capillary detail, they do not adequately reveal the choriocapillaris or other microvascular features beneath the retina. We have developed a noninvasive microvascular imaging technique called phase-variance optical coherence tomography (pvOCT), which identifies vasculature three dimensionally through analysis of data acquired with OCT systems. The pvOCT imaging method is not only capable of generating capillary perfusion maps for the retina, but it can also use the 3D capabilities to segment the data in depth to isolate vasculature in different layers of the retina and choroid. This paper demonstrates some of the capabilities of pvOCT imaging of the anterior layers of choroidal vasculature of a healthy normal eye as well as of eyes with geographic atrophy (GA) secondary to age-related macular degeneration. The pvOCT data presented permit digital segmentation to produce 2D depth-resolved images of the retinal vasculature, the choriocapillaris, and the vessels in Sattler’s and Haller’s layers. Comparisons are presented between en face projections of pvOCT data within the superficial choroid and clinical angiography images for regions of GA. Abnormalities and vascular dropout observed within the choriocapillaris for pvOCT are compared with regional GA progression. The capability of pvOCT imaging of the microvasculature of the choriocapillaris and the anterior choroidal vasculature has the potential to become a unique tool to evaluate therapies and understand the underlying mechanisms of age-related macular degeneration progression.

Noninvasive Imaging of the Foveal Avascular Zone with High-Speed, Phase-Variance Optical Coherence Tomography

PURPOSE To demonstrate the application of phase-variance optical coherence tomography (pvOCT) for contrast agent-free in vivo imaging of volumetric retinal microcirculation in the human foveal region and for extraction of foveal avascular zone dimensions. METHODS A custom-built, high-speed Fourier-domain OCT retinal imaging system was used to image retinas of two healthy subjects and eight diabetic patients. Through the acquisition of multiple B-scans for each scan location, phase differences between consecutive scans were extracted and used for phase-variance contrast, identifying motion signals from within blood vessels and capillaries. The en face projection view of the inner retinal layers segmented out from volumetric pvOCT data sets allowed visualization of a perfusion network with the foveal avascular zone (FAZ). In addition, the authors presented 2D retinal perfusion maps with pseudo color-coded depth positions of capillaries. RESULTS Retinal vascular imaging with pvOCT provides accurate measurements of the FAZ area and its morphology and a volumetric perfusion map of microcapillaries. In this study using two images from each fundus fluorescein angiography (FA) and pvOCT, the measured average areas of the FAZ from two healthy subjects were below 0.22 mm(2), and each of eight diabetic patients had an enlarged FAZ area, larger than 0.22 mm(2). Moreover, the FAZ areas demonstrated a significant correlation (r = 0.91) between measurements from FA and pvOCT. CONCLUSIONS The high-speed pvOCT allows contrast agent-free visualization of capillary networks in the human foveal region that is analogous to fundus FA imaging. This could allow for noninvasive diagnosis and progression monitoring of diabetic retinopathy in clinical settings.

Phase-Contrast OCT Imaging of Transverse Flows in the Mouse Retina and Choroid

PURPOSE To test the hypothesis that a novel phase-contrast optical coherence tomography (OCT) system can image retinal and choroidal vessels in the living mouse. METHODS A high-speed spectral domain optical coherence tomography (SDOCT) system, which measures the reflections for the entire depth of the retina at once with each axial scan (A-scan), was developed for mouse retinal imaging. Acquiring multiple A-scans over a transverse line across the mouse retina offers a two-dimensional cross-sectional image (B-scan); several neighboring B-scans can be assembled into a three-dimensional OCT image. To visualize mobility and transverse flow in retinal vessels, the statistical variance of phase for each location was calculated from multiple B-scans acquired successively for the same retinal cross-section. Such measures of phase variance offer a direct measure of motions over a large dynamic range of flow velocities. RESULTS Three-dimensional phase-contrast images of the live mouse retina were created using multiple two-dimensional cross-sectional image slices through the retina. For the data presented here, each cross-sectional phase-contrast slice resulted from five images of 100 or 200 transverse pixels, acquired over 25 ms or 50 ms, respectively. The approach offered clear identification of motion regions at different depths, including flow in the retinal microvasculature and in the choroidal vessels. CONCLUSIONS Phase-contrast OCT enables three-dimensional visualization of retinal and choroidal vasculature in vivo.

Molecular contrast optical coherence tomography: a pump-probe scheme using indocyanine green as a contrast agent

The use of indocyanine green (ICG), a U.S. Food and Drug Administration approved dye, in a pump-probe scheme for molecular contrast optical coherence tomography (MCOCT) is proposed and demonstrated for the first time. In the proposed pump-probe scheme, an optical coherence tomography (OCT) scan of the sample containing ICG is first acquired. High fluence illumination (approximately 190 kJ/cm2) is then used to permanently photobleach the ICG molecules--resulting in a permanent alteration of the overall absorption of the ICG. A second OCT scan is next acquired. The difference of the two OCT scans is used to determine the depth resolved distribution of ICG within a sample. To characterize the extent of photobleaching in different ICG solutions, we determine the cumulative probability of photobleaching, phi(B,cum), defined as the ratio of the total photobleached ICG molecules to the total photons absorbed by the ground state molecules. An empirical study of ICG photobleaching dynamics shows that phi(B,cum) decreases with fluence as well as with increasing dye concentration. The quantity phi(B,cum) is useful for estimating the extent of photobleaching in an ICG sample (MCOCT contrast) for a given fluence of the pump illumination. The paper also demonstrates ICG-based MCOCT imaging in tissue phantoms as well as within stage 54 Xenopus laevis.

Homodyne en face optical coherence tomography

We demonstrate, for what we believe to be the first time, the use of a 3 x 3 fiber-optic coupler to realize a homodyne optical coherence tomography (OCT) system for en face imaging of highly scattering tissues and turbid media. The homodyne OCT setup exploits the inherent phase shifts between different output ports of a 3 x 3 fiber-optic coupler to extract amplitude information of a sample. Our homodyne en face OCT system features a measured resolution of 14 microm axially and 9.4 microm laterally with a 90 dB signal-to-noise ratio at 10 micros integration time. En face OCT imaging of a stage 52 Xenopus laevis was successfully demonstrated at a depth of 600 microm within the sample.

Phase variance optical coherence microscopy for label-free imaging of the developing vasculature in zebrafish embryos

Abstract. A phase variance optical coherence microscope (pvOCM) has been created to image blood flow in the microvasculature of zebrafish embryos, without the use of exogenous labels. The pvOCM imaging system has axial and lateral resolutions of 2.8  μm in tissue and imaging depth of more than 100  μm. Images of 2 to 5 days postfertilization zebrafish embryos identified the detailed anatomical structure based on OCM intensity contrast. Phase variance contrast offered visualization of blood flow in the arteries, veins, and capillaries. The pvOCM images of the vasculature were confirmed by direct comparisons with fluorescence microscopy images of transgenic embryos in which the vascular endothelium is labeled with green fluorescent protein. The ability of pvOCM to capture activities of regional blood flow permits it to reveal functional information that is of great utility for the study of vascular development.

Visualization of human retinal micro-capillaries with phase contrast high-speed optical coherence tomography

We present high-speed Fourier-domain optical coherence tomography (Fd-OCT) with the phase variance based motion contrast method for visualizing retinal micro-circulation in vivo. This technique allows non-invasive visualization of a two-dimensional retinal perfusion map and concurrent volumetric morphology of retinal microvasculature with high sensitivity. The high-speed acquisition rate at 125kHz A-scans enables reduction of motion artifacts with increased scanning area if compared to previously reported results. Several scanning schemes with different sampling densities and scanning areas are evaluated to find optimal parameters for in vivo imaging. In order to evaluate this technique, we compare OCT micro-capillary imaging using the phase variance technique with fundus fluorescein angiography (FA). Additionally, volumetric visualization of blood flow for a normal subject is presented.