17 February 2017 Lensfree diffractive tomography for the imaging of 3D cell cultures
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Abstract
New microscopes are needed to help reaching the full potential of 3D organoid culture studies by gathering large quantitative and systematic data over extended periods of time while preserving the integrity of the living sample. In order to reconstruct large volumes while preserving the ability to catch every single cell, we propose new imaging platforms based on lens-free microscopy, a technic which is addressing these needs in the context of 2D cell culture, providing label-free and non-phototoxic acquisition of large datasets. We built lens-free diffractive tomography setups performing multi-angle acquisitions of 3D organoid cultures embedded in Matrigel and developed dedicated 3D holographic reconstruction algorithms based on the Fourier diffraction theorem. Nonetheless, holographic setups do not record the phase of the incident wave front and the biological samples in Petri dish strongly limit the angular coverage. These limitations introduce numerous artefacts in the sample reconstruction. We developed several methods to overcome them, such as multi-wavelength imaging or iterative phase retrieval. The most promising technic currently developed is based on a regularised inverse problem approach directly applied on the 3D volume to reconstruct. 3D reconstructions were performed on several complex samples such as 3D networks or spheroids embedded in capsules with large reconstructed volumes up to ~ 25 mm3 while still being able to identify single cells. To our knowledge, this is the first time that such an inverse problem approach is implemented in the context of lens-free diffractive tomography enabling to reconstruct large fully 3D volumes of unstained biological samples.
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Anthony Berdeu, Fabien Momey, Jean-Marc Dinten, Xavier Gidrol, Nathalie Picollet-D'hahan, Cédric Allier, "Lensfree diffractive tomography for the imaging of 3D cell cultures", Proc. SPIE 10070, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXIV, 1007008 (17 February 2017); doi: 10.1117/12.2250152; https://doi.org/10.1117/12.2250152
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