Imaging of living cells based on traditional fluorescence and confocal laser scanning microscopy has delivered an enormous amount of information critical for understanding biological processes in single cells. However, the requirement for a high numerical aperture and fluorescent markers still limits researchers’ ability to visualize the cellular architecture without causing short- and long-term photodamage. Optical coherence microscopy (OCM) is a promising alternative that circumvents the technical limitations of fluorescence imaging techniques and provides unique access to fundamental aspects of early embryonic development, without the requirement for sample pre-processing or labeling. In the present paper, we utilized the internal motion of cytoplasm, as well as custom scanning and signal processing protocols, to effectively reduce the speckle noise typical for standard OCM and enable high-resolution intracellular time-lapse imaging. To test our imaging system we used mouse and pig oocytes and embryos and visualized them through fertilization and the first embryonic division, as well as at selected stages of oogenesis and preimplantation development. Because all morphological and morphokinetic properties recorded by OCM are believed to be biomarkers of oocyte/embryo quality, OCM may represent a new chapter in imaging-based preimplantation embryo diagnostics.
We propose a simple and robust procedure for Fourier domain optical coherence tomography (FdOCT) that allows to linearize the detected FdOCT spectra to wavenumber domain and, at the same time, to determine the wavelength of light for each point of detected spectrum. We show that in this approach it is possible to use any measurable physical quantity that has linear dependency on wavenumber and can be extracted from spectral fringes. The actual values of the measured quantity have no importance for the algorithm and do not need to be known at any stage of the procedure. As example we calibrate a spectral OCT spectrometer using Doppler frequency. The technique of spectral calibration can be in principle adapted to of all kind of Fourier domain OCT devices.
Although Doppler optical coherence tomography techniques have enabled the imaging of blood flow in mid-sized vessels in biological tissues, the generation of velocity maps of capillary networks remains a challenge. To better understand the origin and information content of the Doppler signal from small vessels and limitations of such measurements, we used joint spectral and time domain optical coherence tomography to monitor the flow in a model, semitransparent microchannel device. The results obtained for Intralipid, whole blood, as well as separated red blood cells indicate that the technique is suitable to record velocity profiles in vitro, in a range of microchannel configurations.
We propose a new method and optical instrumentation for mouse brain imaging based on extended-focus optical coherence microscopy. This in vivo imaging technique allows the evaluation of the cytoarchitecture at cellular level and the circulation system dynamics in three dimensions. This minimally invasive and non-contact approach is performed without the application of contrasting agents. The optical design achieved a resolution of 2.2 μm over a distance of 800 μm, which was sufficient to obtain a detailed three-dimensional image of a wild-type mouse's brain down to the layer III of the cortex. Intrinsically contrasted microvessels and structures similar to the bodies of neurons were distinguishable.
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