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Swansea University Medical School

4 ARTICLES PUBLISHED IN JoVE

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Biochemistry

Cell Culture on Silicon Nitride Membranes and Cryopreparation for Synchrotron X-ray Fluorescence Nano-analysis
Caroline Bissardon 1, Solveig Reymond 1, Murielle Salomé 2, Lionel André 2, Sam Bayat 1, Peter Cloetens 2, Sylvain Bohic 1,2
1Inserm, UA7, Synchrotron Radiation for Biomedicine (STROBE), Université Grenoble Alpes, 2ID16A Beamline, ESRF, The European Synchrotron

Presented here is a protocol for cell culture on silicon nitride membranes and plunge-freezing prior to X-ray fluorescence imaging with a synchrotron cryogenic X-ray nanoprobe. When only room temperature nano-analysis is provided, the frozen samples can be further freeze-dried. These are critical steps to obtain information on the intracellular elemental composition.

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Chemistry

Biological Samples Preparation for Speciation at Cryogenic Temperature using High-Resolution X-Ray Absorption Spectroscopy
Caroline Bissardon 1, Marie-Pierre Isaure 2, Emmanuel Lesuisse 3, Mauro Rovezzi 4,5, Eric Lahera 4,5, Olivier Proux 4,5, Sylvain Bohic 1,6
1University Grenoble Alpes, INSERM UA7, Synchrotron Radiation for Biomedicine (STROBE), 2CNRS, UMR 5254, Université Pau & Pays Adour, E2S UPPA, Institut des Sciences Analytiques et de Physicochimie pour l'Environnement et les Matériaux, 3Institut Jacques Monod, UMR 7592 CNRS, Université Paris Diderot, 4OSUG, UMS 832 CNRS, Université Grenoble Alpes, 5FAME and FAME-UHD beamlines, ESRF, the European Synchrotron, 6ID16A beamline, ESRF, the European Synchrotron

This protocol presents a detailed procedure to prepare biological cryosamples for synchrotron-based X-ray absorption spectroscopy experiments. We describe all the steps required to optimize sample preparation and cryopreservation with examples of the protocol with cancer and phytoplankton cells. This method provides a universal standard of sample cryo-preparation.

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Bioengineering

Advanced 3D Liver Models for In vitro Genotoxicity Testing Following Long-Term Nanomaterial Exposure
Samantha V. Llewellyn 1, Gillian E. Conway 1, Ume-Kulsoom Shah 1, Stephen J. Evans 1, Gareth J.S. Jenkins 1, Martin J.D. Clift 1, Shareen H. Doak 1
1In vitro Toxicology Group, Swansea University Medical School

This procedure was established to be used for developing advanced 3D hepatic cultures in vitro, which can provide a more physiologically relevant assessment of the genotoxic hazards associated with nanomaterial exposures over both an acute or long-term, repeated dose regimes.

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Bioengineering

In Vitro Preparation of Actively Maturing Bovine Articular Cartilage Explants for X-Ray Phase Contrast Imaging
Caroline Bissardon 1,2,3, Yadan Zhang 2, Hélène Rougé Labriet 1,4, Sébastien Berujon 5, Laurent Charlet 3, Ilyas M. Khan 2, Emmanuel Brun 1, Sylvain Bohic 1
1Inserm UA07, Rayonnement Synchrotron pour la Recherche Biomédicale (STROBE), 2Centre of Nanohealth, Swansea University Medical School, 3ISTerre, Université Grenoble Alpes, CS, Grenoble Cedex 9, 4Novitom, R&D Grenoble, 5ESRF, European Synchrotron Radiation Facility, CS, Grenoble Cedex 9

This protocol describes the preparation of in vitro bovine articular cartilage for imaging in high resolution with X-rays. These explants actively undergo postnatal maturation. We describe here the necessary steps from the biopsy to data analysis of 3D X-ray phase contrast imaging, passing through explant culture, tissue fixation and synchrotron preparation.

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