Enhancing microvasculature maps for Optical Coherence Tomography Angiography (OCT-A)

Optical coherence microscopy (OCM) is a high-resolution imaging technique based on optical coherence tomography and confocal microscopy. The recent studies on OCM operating at 800-1300 nm spectral region have shown that OCM enables to visualize micrometer- or sub-micrometer-scale structures of animal tissues. Although OCMs offers such high-resolution label-free imaging capability of animal tissues, the imaging depth was restricted by multiple light scattering and light absorption of water in samples. Here, for high-resolution deep-tissue imaging, we developed an OCM in the 1700-nm spectral band by using a supercontinuum (SC) source with a Gaussian-like spectral shape in the wavelength region. Recently, it has been reported that the 1700-nm spectral band is a promising choice for enhancing the imaging depth in the observation of turbid scattering tissues because of the low attenuation coefficient of light. In this study, to clarify that the 1700-nm OCM has a potential to realize the enhanced imaging depth, we compared the attenuation of the signal-to-noise ratio between the 1700-nm and 1300-nm OCM imaging of a mouse brain under the same signal detection sensitivity condition. The result shows that the 1700-nm OCM enables us to achieve the enhanced imaging depth. In this 1700-nm OCM, we also confirmed that the lateral resolution of 1.3 µm and axial resolution of 2.8 µm in tissue were achieved.

Optical coherence tomography (OCT) is a non-invasive cross-sectional imaging technique with micrometer resolution. OCT is useful, non-invasive imaging technique of the internal structure, and it has been applied in many fields, especially medicine and industry. The theoretical axial resolution is determined by the center wavelength and bandwidth of the light source, and the wider the bandwidth is, the higher the axial resolution is. Supercontinuum is the high-power, ultrawideband light source. We have been investigating ultrahigh-resolution (UHR)-OCT using supercontinuum. The characteristics of OCT imaging depend on the optical wavelength used. In this talk, we report our recent work of the wavelength dependence of UHR-OCT using a supercontinuum for biomedical imaging. In order to investigate the wavelength dependence of UHR-OCT, the wideband, high-power, low-noise supercontinua were generated at wavelengths of 0.8, 1.1, 1.3, and 1.7 um based on ultrashort pulses and nonlinear fibers. The wavelength dependence of OCT imaging was examined quantitatively using biological phantoms. Ultrahigh-resolution imaging of a rat lung was demonstrated with wavelengths of 0.8 – 1.0 um UHR-OCT. The variation of alveolar volume was estimated using 3D image analysis. We also developed UHR-spectral domain-OCT and optical coherence microscopy (OCM) at 1.7 um. The high-resolution and high-penetration imaging of turbid tissue, especially mouse brain, was demonstrated. The wavelength dependence of OCM was also discussed in terms of mouse brain imaging.

Optical coherence tomography (OCT) is a non-invasive cross-sectional imaging technique with micrometer resolution. We have been investigating ultrahigh-resolution (UHR)-OCT using supercontinuum. The characteristics of OCT imaging depend on the optical wavelength used. In order to investigate the wavelength dependence of UHR-OCT, the wideband, high-power, low-noise supercontinua were generated at wavelengths of 0.8, 1.1, 1.3, and 1.7 um based on ultrashort pulses and nonlinear fibers. The wavelength dependence of OCT imaging was examined quantitatively using biological phantoms and rat lung tissue. Then we developed UHR-OCT and optical coherence microscopy (OCM) at 1.7 um, which is called as “Optical Window III”. The high-resolution and high-penetration imaging of mouse brain was demonstrated.

Optical coherence microscopy (OCM) has unique advantages of high-resolution volumetric imaging without relying on exogenous labels or dyes. It combines the coherence-gated depth discrimination of optical coherence tomography (OCT) with the high lateral resolution of confocal microscopy, offering an excellent balance between the resolutions and imaging depth. However, as the lateral resolution becomes higher, the imaging depth of OCM decreases and its three-dimensional imaging capability is greatly degraded. To overcome this limitation, we used amplitude apodization to create quasi-Bessel beam illumination in order to extend the depth of focus. The lateral and axial resolutions of our OCM system were measured to be 1.6 μm and 2.9 μm in tissue. The imaging depth was extended by ~3.0X (~100 μm) beyond that of the standard Gaussian beam OCM. Using zebrafish embryos as a test system, we demonstrate extendedfocus OCM for structural imaging studies, which revealed the detailed anatomy deep in embryos.

Towards high resolution imaging of human airways with optical coherence microscopy

Optical coherence microscopy (OCM) is a non-destructive 3-dimentional (3D) imaging technique with micrometer-scale resolution, which is based on optical coherence tomography (OCT) and confocal microscopy [1-3]. Recently, it has been demonstrated that the OCT imaging depth is improved by using 1700 nm spectral band, so called “optical window 3”, due to the lower light absorption and a local minimum of water absorption in the wavelength region [4-8]. In our group, we developed a supercontinuum (SC) laser source at the 1700-nm wavelength region and demonstrated that 1700-nm OCT allows us to visualize the structures of mouse brain at a depth up to 1.7 mm [9-11]. We also successfully demonstrated 3D highresolution visualization of myelinated axons in a mouse brain with 1700-nm OCM, which we developed based on a time-domain low-coherence interferometer [12]. In this presentation, we report high-resolution spectral-domain OCM (SD-OCM) at the 1700-nm spectral band for improving the imaging acquisition speed of 1700-nm OCM.

Optical coherence microscopy (OCM) is a novel technique complementary to optical coherence tomography (OCT) which combines low-coherence interferometry with confocal microscopy to achieve micron-scale resolution imaging in highly scattering media. OCM may be implemented using a single-mode fiber-optic low-coherence interferometer (See Fig. 1). A high numerical aperture objective is used to focus sample-arm light into the specimen, and the reference arm length of the interferometer is adjusted to match the sample arm focal plane optical depth. The sample arm of the interferometer comprises a scanning confocal microscope, in which either the sample or the probe beam is laterally scanned in a raster pattern, and the optical fiber acts as a single-mode confocal aperture for combined light illumination and collection. The reference arm length of the interferometer establishes the depth position of an interferometric “coherence gate” in the sample, from which backscattered light is preferentially collected. Initial studies of OCM in scattering phantoms have demonstrated that this technique provides increased optical sectioning depth compared to confocal microscopy alone.

To investigate optical coherence microscopy (OCM) imaging features and the application value of these high-resolution images for identifying endocervical canal lesions (ECLs), which is a clinical dilemma in cervical cancer screening programs. In total, 520 OCM images were obtained by scanning the cervical canal lesions with an ultra-high-resolution OCM system (204 specimens from 73 patients). The OCM morphologic characteristics of ECLs were observed and summarized, and then 3 researchers performed a diagnostic test of OCM images of cervical canal lesions. The accuracy, sensitivity, specificity, positive predictive value, negative predictive value, 95% confidence interval of each parameter, and interinvestigator agreement (κ) were calculated. Normal endocervix, cysts, squamous metaplasia, high-grade squamous intraepithelial lesions involving glands, and invasive carcinoma had distinct OCM characteristics, which correlated well with corresponding H&E histologic sections. The accuracy, sensitivity, and specificity of the 3 researchers were 90.6%, 89.3% (95% CI, 86.5%-91.7%) and 91.6% (95% CI, 89.2%-93.5%), respectively. The positive predictive value was 90.1% (95% CI, 87.3%-92.4%), and the negative predictive value was 90.9% (95% CI, 88.5%-92.9%), with almost perfect agreement (κ = 0.874). The application of the OCM system in cervical canal lesions is feasible and could help improve detection of occult ECLs in cervical cancer screening programs. This study lays the foundation for further research on OCM in cervical canal lesions in vivo, which also has a potential impact on projecting pathologic evaluation beyond what is currently possible, perhaps globally.

We quantitatively investigated the image quality in deep tissue imaging with optical coherence microscopy (OCM) in the 1700 nm spectral band, in terms of the signal-to-background ratio (SBR) and lateral resolution. In this work, to demonstrate the benefits of using the 1700 nm spectral band for OCM imaging of brain samples, we compared the imaging quality of OCM en-face images obtained at the same position by using a hybrid 1300 nm/1700 nm spectral domain (SD) OCM system with shared sample and reference arms. By observing a reflective resolution test target through a 1.5 mm-thick tissue phantom, which had a similar scattering coefficient to brain cortex tissue, we confirmed that 1700 nm OCM achieved an SBR about 6-times higher than 1300 nm OCM, although the lateral resolution of the both OCMs was similarly degraded with the increase of the imaging depth. Finally, we also demonstrated high-contrast deep tissue imaging of a mouse brain at a depth up to 1.8 mm by using high-resolution 1700 nm SD-OCM.

Optical coherence tomography (OCT) is a noninvasive cross-sectional imaging technique with micrometer resolution. The theoretical axial resolution is determined by the center wavelength and bandwidth of the light source, and the wider the bandwidth, the higher the axial resolution. The characteristics of OCT imaging depend on the optical wavelength used. In this paper, we investigated the wavelength dependence of ultrahigh-resolution (UHR) OCT using a supercontinuum for biomedical imaging. Wideband, high-power, low-noise supercontinua (SC) were generated at λ = 0.8, 1.1, 1.3, and 1.7 μ m based on ultrashort pulses and nonlinear fibers. The wavelength dependence of OCT imaging was examined quantitatively using biological phantoms. Ultrahigh-resolution imaging of a rat lung was demonstrated with λ = 0.8–1.0 μ m UHR-OCT. The variation of alveolar volume was estimated using three-dimensional image analysis. Finally, UHR-spectral domain-OCT and optical coherence microscopy at 1.7 μ m were developed, and high-resolution and high-penetration imaging of turbid tissue, especially mouse brain, was demonstrated.

.We present a multimodal visible light optical coherence microscopy (OCM) and fluorescence imaging (FI) setup. Specification and phantom measurements were performed to characterize the system. Two applications in neuroimaging were investigated. First, curcumin-stained brain slices of a mouse model of Alzheimer’s disease were examined. Amyloid-beta plaques were identified based on the fluorescence of curcumin, and coregistered morphological images of the brain tissue were provided by the OCM channel. Second, human brain tumor biopsies retrieved intraoperatively were imaged prior to conventional neuropathologic work-up. OCM revealed the three-dimensional structure of the brain parenchyma, and FI added the tumor tissue-specific contrast. Attenuation coefficients computed from the OCM data and the florescence intensity values were analyzed and showed a statistically significant difference for 5-aminolevulinic acid (5-ALA)-positive and -negative brain tissues. OCM findings correlated well with malignant hot spots within brain tumor biopsies upon histopathology. The combination of OCM and FI seems to be a promising optical imaging modality providing complementary contrast for applications in the field of neuroimaging.

We demonstrate methods for achieving high resolution imaging using alternate scanning techniques in optical coherence tomography and optical coherence microscopy. These techniques enable high transverse resolutions and overcome depth of field limitations. Cellular level resolutions in human tissue may be achieved.

First experimental results on OCT imaging of internal structure of plant tissues and in situ OCT monitoring of plant tissue regeneration at different water supply are reported. Experiments for evaluating OCT capabilities were performed on Tradescantia. The investigation of seeds swelling was performed on wheat seeds (Triticum L.), barley seeds (Hordeum L.), long-fibred flax seeds (Linum usitatissimum L.) and cucumber seeds (Cucumis sativus L.). These OCT images correlate with standard microscopy data from the same tissue regions. Seeds were exposed to a low-intensity physical factor-the pulsed gradient magnetic field (GMF) with pulse duration 0.1 s and maximum amplitude 5 mT (4 successive pulses during 0.4 s). OCT and OCM enable effective monitoring of fast reactions in plants and seeds at different water supply.

The murine model is a common model for studying developmental diseases. In this study, we compare the performance of the relatively new method of Optical Projection Tomography (OPT) to the well-established technique of Optical Coherence Tomography (OCT) to assess murine embryonic development at three stages, 9.5, 11.5, and 13.5 days post conception. While both methods can provide spatial resolution at the micrometer scale, OPT can provide superior imaging depth compared to OCT. However, OPT requires samples to be fixed, placed in an immobilization media such as agar, and cleared before imaging. Because OCT does not require fixing, it can be used to image embryos in vivo and in utero. In this study, we compare the efficacy of OPT and OCT for imaging murine embryonic development. The data demonstrate the superior capability of OPT for imaging fine structures with high resolution in optically-cleared embryos while only OCT can provide structural and functional imaging of live embryos ex vivo and in utero with micrometer scale resolution.

The mouse provides an excellent model for studying mammalian development and disease because of the resemblance to humans, potential for genetic manipulation, and rapid generation time. However, mammalian embryos develop in utero, which limits the ability to observe development with high-resolution. In this study, we adopted Swept Source Optical Coherence Tomography (SS-OCT) for live visualization of mouse embryos during different stages of the development. We demonstrate capability of OCT as a novel methodology for live imaging of mouse embryos both in culture and through the uterine wall.