KEYWORDS: Optical coherence tomography, Skin, In vitro testing, Temperature metrology, Mode conditioning cables, Signal processing, Scattering, Acquisition tracking and pointing, 3D modeling, Motion models
Microscopic optical coherence tomography (OCT) provides three-dimensional, high-resolution imaging but lacks (sub-) cellular contrast. Dynamic-microscopic OCT (dmOCT) is an approach exploiting dynamic changes of the scattering behavior in metabolically active cells. However, the underlying cellular processes responsible for those intensity fluctuations and hence the dynamic signals are not finally identified yet. Here, we present the effects of different temperatures and metabolic reagents on dmOCT images of an in-vitro human skin model. Our data indicates a dependency of the dmOCT signals on metabolic activity rather than Brownian motion and suggests dependency on the metabolic state.
Optical coherence tomography (OCT) with axial and lateral resolution of 1 µm, termed microscopic OCT (mOCT), is suited for the investigation of dynamic processes on cellular level. An improvement in contrast to visualize cellular structures can be achieved by evaluating inherent signal fluctuation. This so-called dynamic contrast was recently demonstrated for the widely used scanning frequency domain OCT (FD-OCT). Here we show comparative measurements of dynamic microscopic OCT (dmOCT) and multiphoton autofluorescence imaging of ex-vivo trachea. Compared to multiphoton microscopy, the dmOCT provides greater penetration depth, can visualize structures that are not detectable by autofluorescence imaging and has no risk of photodamage.
Here we present a forward-looking endoscope for dynamic microscopic OCT reaching a lateral resolution of 1.3 µm and 0.8 mm field of view. Since tissue motion degrades dynamic imaging, tissue was immobilized by suction. The endoscope was placed in a 4 mm stainless-steel sheath, which was connected to a vacuum pump. In mice, the endoscope can access various inner organs using open surgery or laparoscopy. The potential of the dynamic endo-microscopic OCT was demonstrated on relevant murine tissue such as liver, spleen and kidney. Otherwise invisible cellular and subcellular structures were imaged by dynamic mOCT with high contrast.
Optical coherence tomography (OCT) with microscopic resolution is suited for the investigation of dynamic processes on cellular level. Most existing setups for microscopic OCT (mOCT) are able to acquire A-Scans at 100 kHz, hence they are suitable for displaying only B-Scans in real-time. We present an mOCT setup with a new high-speed spectrometer, which is capable of acquiring up to 600.000 A-scans/s. Customized software allows to image mOCT volumes with micrometer resolution at video rate. Therefore, we are able to visualize functional processes on a cellular level in three dimensions over time. Here, we present 2D and 4D data of ex vivo human lung. Recording tens of high-resolution OCT volumes per seconds becomes possible faster than previous setup for OCT with microscopic resolution.
Optical coherence tomography (OCT) images scattering tissues with 5 to 15 μm resolution. This is usually not sufficient for a distinction of cellular and subcellular structures. Increasing axial and lateral resolution and compensation of artifacts caused by dispersion and aberrations is required to achieve cellular and subcellular resolution. This includes defocus which limit the usable depth of field at high lateral resolution. OCT gives access the phase of the scattered light and hence correction of dispersion and aberrations is possible by numerical algorithms. Here we present a unified dispersion/aberration correction which is based on a polynomial parameterization of the phase error and an optimization of the image quality using Shannon’s entropy. For validation, a supercontinuum light sources and a costume-made spectrometer with 400 nm bandwidth were combined with a high NA microscope objective in a setup for tissue and small animal imaging. Using this setup and computation corrections, volumetric imaging at 1.5 μm resolution is possible. Cellular and near cellular resolution is demonstrated in porcine cornea and the drosophila larva, when computational correction of dispersion and aberrations is used. Due to the excellent correction of the used microscope objective, defocus was the main contribution to the aberrations. In addition, higher aberrations caused by the sample itself were successfully corrected. Dispersion and aberrations are closely related artifacts in microscopic OCT imaging. Hence they can be corrected in the same way by optimization of the image quality. This way microscopic resolution is easily achieved in OCT imaging of static biological tissues.
A holographic method for high-speed, noncontact photoacoustic tomography is introduced and evaluated. Relative changes of the object’s topography, induced by the impact of thermoelastic pressure waves, were determined at nanometer sensitivity without physical contact. The object’s surface was illuminated with nanosecond laser pulses and imaged with a high-speed CMOS camera. From two interferograms measured before and after excitation of the acoustic wave, surface displacement was calculated and then used as the basis for a tomographic reconstruction of the initial pressure caused by optical absorption. The holographic detection scheme enables variable sampling rates of the photoacoustic signal of up to 50 MHz. The total acquisition times for complete volumes with 230 MVoxel is far below 1 s. Measurements of silicone and porcine skin tissue phantoms with embedded artificial absorbers, which served as a model for human subcutaneous vascular networks, were possible. Three-dimensional reconstructions of the absorbing structures show details with a diameter of 310 μm up to a depth of 2.5 mm. Theoretical limitations and the experimental sensitivity, as well as the potential for in vivo imaging depending on the detection repetition rate, are analyzed and discussed.
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