Holo-Tomographic Flow Cytometry is a new technology for single-cell analysis that combine Phase-Contrast Tomography and Flow Cytometry opening to a new approach in biomedical field by high-throughput, tri-dimensional imaging of unstained cell populations. Tomographic Phase Microscopy is a label-free phase contrast imaging method able to supply quantitative and volumetric refractive index distribution at single cell level in adherent or fixed populations. Here, we demonstrate that phase-contrast tomography can be achieved also for cells into a microfluidic environment obtaining accurate 3D tomographic imaging of thousands of flowing and rotating cells thanks to a robust and reliable computational strategy. Recording setups are based on Digital Holography in microscopy configurations integrated with microfluidic apparatus to record interference fringes (hologram) of rotating cells. Computational pipeline includes 3D cell tracking into the microfluidic channel, quantitative 2D phase-contrast maps retrieval for each acquired hologram, robust angle recovery code, tomographic processing to measure the inner refractive index distribution. Holo-Tomographic Flow Cytometry surpasses the limits of conventional Imaging flow cytometers because make available the recording of hundreds of informative images for each flowing cells avoiding the employment of fluorescent tags. Holo-Tomographic Flow Cytometry allows to retrieve the unique all-optical 3D fingerprint for each cell flowing into the field-of-view opening to a wide range of applications such as: (i) identification of inner subcellular compartments; (ii) recognition of nanoparticle uptake and (iii) phenotyping of different subclasses in heterogeneous populations. Future perspectives are presented in the fields of liquid biopsy, drug resistance and genetic disfunctions.
Human health and disease prevention are among the priorities to safeguard astronauts and, in the next future, space tourists. There is a great demand of new reliable biotechnologies that would be eventually implemented on spacecrafts to observe the space-induced effect on humans. One of the main risks is related to the radiation exposure, that is significantly higher than on Earth. For this reason, space agencies are pushing to develop strategies to quantify, oversee and limiting such risks. Here we present an approach based on the combination of microfluidics and stain-free imaging also aided by artificial intelligence to monitor the effect on ionizing radiation on blood cells. The system is based on the Holographic Image Flow Cytometry system where Quantitative Phase Contrast images are retrieved for cell flowing and rotating into a microfluidics circuits. Proof of concept is demonstrated where morphological parameters are identified able to distinguish cell population irradiated at different radiation doses and at different time from the radiation exposure. Blood cell will be analyzed. The presented approach has main advantages respect to standard and already existent technologies for single cell analysis. The first one is the no-need of fluorescence staining thus opening to faster and easier operation steps. The second one is related to cell rotation into the field of view, allowing to acquire images at different rotation angle and thus collecting a broader dataset useful for the application of artificial intelligence network. Furthermore, the system can be miniaturized to a scale portable out of the laboratory environment.
The interaction between light and biological matter can be exploited as a useful tool in various fields of science and technology. Indeed the optical behavior of living cells can permit to use them as micro-lenses for imaging, as photonic micro-resonators or waveguides, and also as advanced probes in holographic optical tweezers for manipulating the matter at nanoscale, and even bio-probes of localized fluorescence at sub-wavelength scale have been demonstrated. Here, we present an overview of these new insights about biological lenses. Theoretical modelling of the lensing effect of living cells will be discussed in details in case of Red Blood Cells. Digital holography in microscopy configuration is the tool that allow the experimental verification of this modelling thanks to the numerical refocusing capability. Applications of such new paradigm range from anemia diagnostics to bio-lithography.
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