Ultrahigh-resolution imaging of human donor cornea using full-field optical coherence tomography

A feasibility study of ultrahigh-resolution full-field optical coherence tomography (FF-OCT) for a subcellular-level imaging of human donor corneas is presented. The FF-OCT system employed in this experiment is based on a white light interference microscope, where the sample is illuminated by a thermal light source and a horizontal cross-sectional (en face) image is detected using a charge coupled device (CCD) camera. A conventional four-frame phase-shift detection technique is employed to extract the interferometric image from the CCD output. A 95-nm-broadband full-field illumination yields an axial resolution of 2.0 microm, and the system covers an area of 850 microm x 850 microm with a transverse resolution of 2.4 microm using a 0.3-NA microscope objective and a CCD camera with 512 x 512 pixels. Starting a measurement from the epithelial to the endothelial side, a series of en face images was obtained. From detected en face images, the epithelial cells, Bowman's layer, stromal keratocyte, nerve fiber, Descemet's membrane, and endothelial cell were clearly observed. Keratocyte cytoplasm, its nuclei, and its processes were also separately detected. Two-dimensional interconnectivity of the keratocytes is visualized, and the keratocytes existing between collagen lamellaes are separately extracted by exploiting a high axial resolution ability of FF-OCT.     

In vivo human retinal imaging by Fourier domain optical coherence tomography

We present what is to our knowledge the first in vivo tomograms of human retina obtained by Fourier domain optical coherence tomography. We would like to show that this technique might be as powerful as other optical coherence tomography techniques in the ophthalmologic imaging field. The method, experimental setup, data processing, and images are discussed.     

High-resolution imaging diagnosis of human fetal membrane by three-dimensional optical coherence tomography

Microscopic chorionic pseudocyst (MCP) arising in the chorion leave of the human fetal membrane (FM) is a clinical precursor for preeclampsia which may progress to fatal medical conditions (e.g., abortion) if left untreated. To examine the utility of three-dimensional (3D) optical coherence tomography (OCT) for noninvasive delineation of the morphology of human fetal membranes and early clinical detection of MCP, 60 human FM specimens were acquired from 10 different subjects undergoing term cesarean delivery for an ex vivo feasibility study. Our results showed that OCT was able to identify the four-layer architectures of human FMs consisting of high-scattering decidua vera (DV, average thickness d(DV) ≈ 92±38 μm), low-scattering chorion and trophoblast (CT, d(CT) ≈ 150±67 μm), high-scattering subepithelial amnion (A, d(A) ≈ 95±36 μm), and low-scattering epithelium (E, d(E) ≈ 29±8 μm). Importantly, 3D OCT was able to instantaneously detect MCPs (low scattering due to edema, fluid buildup, vasodilatation) and track (staging) their thicknesses d(MCP) ranging from 24 to 615 μm. It was also shown that high-frequency ultrasound was able to compliment OCT for detecting more advanced thicker MCPs (e.g., d(MCP)>615 μm) because of its increased imaging depth.     

In vivo imaging of human burn injuries with polarization-sensitive optical coherence tomography

The accurate determination of burn depth is critical in the clinical management of burn wounds. Polarization-sensitive optical coherence tomography (PS-OCT) has been proposed as a potentially non-invasive method for determining burn depth by measuring thermally induced changes in the structure and birefringence of skin, and has been investigated in pre-clinical burn studies with animal models and ex vivo human skin. In this study, we applied PS-OCT to the in-vivo imaging of two pediatric burn patients. Deep and superficial burned skins along with contralateral controls were imaged in 3D. The imaging size was 8 mm × 6 mm × 2 mm in width, length, and depth in the air respectively, and the imaging time was approximately 6 s per volume. Superficially burned skins exhibited the same layered structure as the contralateral controls, but more visible vasculature and reduced birefringence compared to the contralateral controls. In contrast, a deeply burned skin showed loss of the layered structure, almost absent vasculature, and smaller birefringence compared to superficial burns. This study suggested the vasculature and birefringence as parameters for characterizing burn wounds.     

Comparison of optical coherence tomographic assessment between first- and second-generation drug-eluting stents

Purpose

There is a lack of sufficient data in comparison of optical coherence tomographic (OCT) findings between first- and second-generation drug-eluting stents (DES). Compared to first-generation (i.e., sirolimus- or paclitaxel-eluting stents), second-generation DESs (i.e., everolimus- or biolinx-based zotarolimus-eluting stents) might have more favorable neointimal coverage.

Materials and Methods

Follow-up OCT findings of 103 patients (119 lesions) treated with second-generation DESs were compared with those of 139 patients (149 lesions) treated with first-generation DESs. The percentage of uncovered or malapposed struts, calculated as the ratio of uncovered or malapposed struts to total struts in all OCT cross-sections, respectively, was compared between the two groups.

Results

Both DES groups showed similar suppression of neointimal hyperplasia (NIH) on OCT (mean NIH cross-sectional area; second- vs. first-generation=1.1±0.5 versus 1.2±1.0 mm², respectively, p=0.547). However, the percentage of uncovered struts of second-generation DESs was significantly smaller than that of first-generation DESs (3.8±4.8% vs.7.5±11.1%, respectively, p<0.001). The percentage of malapposed struts was also significantly smaller in second-generation DESs than in first-generation DESs (0.4±1.6% vs.1.4±3.7%, respectively, p=0.005). In addition, intra-stent thrombi were less frequently detected in second-generations DESs than in first-generation DESs (8% vs. 20%, respectively, p=0.004).

Conclusion

This follow-up OCT study showed that second-generation DESs characteristically had greater neointimal coverage than first-generation DESs.     

Subcellular imaging of epithelium with time-lapse optical coherence tomography

We present the first experimental result of direct delineation of the nuclei of living rat bladder epithelium with ultrahigh-resolution optical coherence tomography (uOCT). We demonstrate that the cellular details embedded in the speckle noise in a uOCT image can be uncovered by time-lapse frame averaging that takes advantage of the micromotion in living biological tissue. The uOCT measurement of the nuclear size (7.9+/-1.4 microm) closely matches the histological evaluation (7.2+/-0.8 microm). Unlike optical coherence microscopy (OCM), which requires a sophisticated high-NA microscopic objective, this approach uses a commercial-grade single achromatic lens (f/10 mm, NA/0.25) and provides a cross-sectional image over 0.6 mm of depth without focus tracking, thus holding great promise of endoscopic optical biopsy for diagnosis and grading of flat epithelial cancer such as carcinoma in situ in vivo.     

High-resolution optical coherence tomography imaging of the living kidney

Optical coherence tomography (OCT) is a rapidly emerging imaging modality that can provide non-invasive, cross-sectional, high-resolution images of tissue morphology in situ and in real-time. In the present series of studies, we used a high-speed OCT imaging system equipped with a frequency-swept laser light source (1.3 mum wavelength) to study living kidneys in situ. Adult, male Munich-Wistar rats were anesthetized, a laparotomy was performed and the living kidneys were exposed for in situ observation. We observed the kidneys prior to, during and following exposure to renal ischemia induced by clamping the renal artery. The effects of intravenous mannitol infusion (1.0 ml of 25%) prior to and during renal ischemia were also studied. Finally, living kidneys were flushed with a renal preservation solution, excised and observed while being stored at 0-4 degrees C. Three-dimensional OCT data sets enabled visualization of the morphology of the uriniferous tubules and the renal corpuscles. When renal ischemia was induced, OCT revealed dramatic shrinkage of tubular lumens due to swelling of the lining epithelium. Three-dimensional visualization and volumetric rendering software provided an accurate evaluation of volumetric changes in tubular lumens in response to renal ischemia. Observations of kidneys flushed with a renal preservation solution and stored at 0-4 degrees C also revealed progressive and significant loss of tubular integrity over time. Intravenous infusion of mannitol solution resulted in thinning of the tubular walls and an increase in the tubular lumen diameters. Mannitol infusion also prevented the cell swelling that otherwise resulted in shrinkage of proximal tubule lumens during ischemia. We conclude that OCT represents an exciting new approach to visualize, in real-time, pathological changes in the living kidney in a non-invasive fashion. Possible clinical applications are discussed.     

Subgingival calculus imaging based on swept-source optical coherence tomography

We characterized and imaged dental calculus using swept-source optical coherence tomography (SS-OCT). The refractive indices of enamel, dentin, cementum, and calculus were measured as 1.625 ± 0.024, 1.534 ± 0.029, 1.570 ± 0.021, and 2.097 ± 0.094, respectively. Dental calculus leads strong scattering properties, and thus, the region can be identified from enamel with SS-OCT imaging. An extracted human tooth with calculus is covered with gingiva tissue as an in vitro sample for tomographic imaging.     

Enhanced optical clearing of skin in vivo and optical coherence tomography in-depth imaging

The strong optical scattering of skin tissue makes it very difficult for optical coherence tomography (OCT) to achieve deep imaging in skin. Significant optical clearing of in vivo rat skin sites was achieved within 15 min by topical application of an optical clearing agent PEG-400, a chemical enhancer (thiazone or propanediol), and physical massage. Only when all three components were applied together could a 15 min treatment achieve a three fold increase in the OCT reflectance from a 300 μm depth and 31% enhancement in image depth Z(threshold).     

Jones-matrix imaging of biological tissues with quadruple-channel optical coherence tomography

Two-dimensional depth-resolved Jones-matrix images of scattering biological tissues were measured with novel double-source double-detector polarization-sensitive optical coherence tomography (OCT). The Jones matrix can be determined in a single scan with this OCT system. The experimental results show that this system can be effectively applied to the measurement of soft tissues, which are less stable than hard tissues. Polarization parameters such as diattenuation, birefringence, and orientation of the fast axis can be extracted through decomposition of the measured Jones matrix. The Jones matrix of thermally treated porcine tendon showed a reduction of birefringence from thermal damage. The Jones matrices of porcine skin and bovine cartilage also revealed that the density and orientation of the collagen fibers in porcine skin and bovine cartilage are not distributed as uniformly as in porcine tendon. Birefringence is sensitive to changes in tissue because it is based on phase contrast.     

High-resolution three-dimensional imaging of biofilm development using optical coherence tomography

We describe the use of optical coherence tomography (OCT) for high-resolution, real-time imaging of three-dimensional structure and development of a Pseudomonas aeruginosa biofilm in a standard capillary flow-cell model. As the penetration depth of OCT can reach several millimeters in scattering samples, we are able to observe complete biofilm development on all surfaces of a 1 mm x 1 mm flow-cell. We find that biofilm growing at the bottom of the tube has more structural features including voids, outward projections, and microcolonies while the biofilm growing on the top of the tube is relatively flat and contains less structural features. Volume-rendered reconstructions of cross-sectional OCT images also reveal three-dimensional structural information. These three-dimensional OCT images are visually similar to biofilm images obtained with confocal laser scanning microscopy, but are obtained at greater depths. Based on the imaging capabilities of OCT and the biofilm imaging data obtained, OCT has potential to be used as a non-invasive, label-free, real-time, in-situ and/or in-vivo imaging modality for biofilm characterization.     

Measurement and imaging of birefringent properties of the human cornea with phase-resolved, polarization-sensitive optical coherence tomography

Optical coherence tomography (OCT) is an emerging technology for high-resolution, noncontact imaging of transparent and scattering media. Polarization-sensitive optical coherence tomography (PS-OCT) is a functional extension of OCT that can image birefringent properties of a biological sample. PS-OCT was used to measure and image retardation and birefringent axis orientation of in vitro human cornea. We used a two-channel PS-OCT system employing a phase-sensitive recording of the interferometric signals in two orthogonal polarization channels. Using an algorithm based on a Hilbert transform, it is possible to calculate the retardation and the slow axis orientation of the sample with only a single A-scan per transversal measurement location. While the retardation information is encoded in the amplitude ratio of the two interferometric signals, the axis orientation is encoded entirely in their phase difference. We present maps of retardation and the distribution of slow axis orientation of the human cornea in longitudinal cross-sections and en face images obtained at the back surface of the cornea. The retardation increases in a radial direction and with depth; the slow axis varies in the transversal direction. Knowledge of the retardation and the slow axis distribution of the cornea might improve nerve fiber polarimetry for glaucoma diagnostics and could be useful for diagnosing different types of pathologies of the cornea.     

High-resolution three-dimensional optical coherence tomography imaging of kidney microanatomy ex vivo

Optical coherence tomography (OCT) is an emerging medical imaging technology that enables high-resolution, noninvasive, cross-sectional imaging of microstructure in biological tissues in situ and in real time. When combined with small-diameter catheters or needle probes, OCT offers a practical tool for the minimally invasive imaging of living tissue morphology. We evaluate the ability of OCT to image normal kidneys and discriminate pathological changes in kidney structure. Both control and experimental preserved rat kidneys were examined ex vivo by using a high-resolution OCT imaging system equipped with a laser light source at 1.3-microm wavelength. This system has a resolution of 3.3 microm (depth) by 6 microm (transverse). OCT imaging produced cross-sectional and en face images that revealed the sizes and shapes of the uriniferous tubules and renal corpuscles. OCT data revealed significant changes in the uriniferous tubules of kidneys preserved following an ischemic or toxic (i.e., mercuric chloride) insult. OCT data was also rendered to produce informative three-dimensional (3-D) images of uriniferous tubules and renal corpuscles. The foregoing observations suggest that OCT can be a useful non-excisional, real-time modality for imaging pathological changes in donor kidney morphology prior to transplantation.     

Numerical analysis of gradient index lens-based optical coherence tomography imaging probes

We report the numerical analysis of gradient index (GRIN) lens-based optical coherence tomography imaging probes to derive optimal design parameters. Long and short working distance probes with a small focal spot are considered. In each model, the working distance and beam waist are characterized and compared for different values of length and refractive index of the probe components. We also explore the influence of the outer tubing and refractive index of the sample media. Numerical results show that the adjustment of the maximum beam diameter and focusing angle at the end of the GRIN lens surface is very important for determining the optical performance parameters of the probe.     

Polarization memory effect in optical coherence tomography and dental imaging application

We report the existence of polarization memory effect (PME) in optical coherence tomography and investigate its potential applications in dental imaging. We performed the study in three steps. First, microsphere scattering phantoms of different sizes were imaged in order to validate experimental results with PME theory. Both linearly and circularly polarized light were used to probe the samples. Second, healthy tooth samples were scanned and polarization memory effect was identified in dentin. In this step, specific verification and signal processing were performed to rule out possible image interpretation by birefringence effect. Third, we evaluated dentin demineralization with PME. Results show polarization memory can be useful to characterize this dynamic mineralization process for early caries detection and rehabilitation.     

Broadband superluminescent diode-based ultrahigh resolution optical coherence tomography for ophthalmic imaging

Spectral domain optical coherence tomography (SD-OCT) with ultrahigh resolution can be used to measure precise structures in the context of ophthalmic imaging. We designed an ultrahigh resolution SD-OCT system based on broadband superluminescent diode (SLD) as the light source. An axial resolution of 2.2 μm in tissue, a scan depth of 1.48 mm, and a high sensitivity of 93 dB were achieved by the spectrometer designed. The ultrahigh-resolution SD-OCT system was employed to image the human cornea and retina with a cross-section image of 2048 × 2048 pixels. Our research demonstrated that ultrahigh -resolution SD-OCT can be achieved using broadband SLD in a simple way.