Université Grenoble Alpes

7 ARTICLES PUBLISHED IN JoVE

Environment

Dispersion of Nanomaterials in Aqueous Media: Towards Protocol Optimization

Inder Kaur¹, Laura-Jayne Ellis¹, Isabella Romer¹, Ratna Tantra², Marie Carriere^3,4, Soline Allard⁵, Martine Mayne-L'Hermite⁵, Caterina Minelli⁶, Wolfgang Unger⁷, Annegret Potthoff⁸, Steffi Rades⁷, Eugenia Valsami-Jones¹

¹School of Geography, Earth and Environmental Sciences, University of Birmingham, ²Analytical Science, National Physical Laboratory, ³INAC-LCIB, Université Grenoble Alpes, ⁴CEA, INAC-SyMMES, ⁵NIMBE, CEA, CNRS, Université Paris-Saclay, CEA Saclay, ⁶Chemical, Medical and Environmental Science, National Physical Laboratory, ⁷BAM Division 6.1 'Surface Analysis and Interfacial Chemistry', BAM Federal Institute for Materials Research and Testing, ⁸Fraunhofer Institute for Ceramic Technologies and Systems

Here, we present a step-wise protocol for the dispersion of nanomaterials in aqueous media with real-time characterization to identify the optimal sonication conditions, intensity, and duration for improved stability and uniformity of nanoparticle dispersions without impacting the sample integrity.

Biology

Lens-free Video Microscopy for the Dynamic and Quantitative Analysis of Adherent Cell Culture

Cedric Allier¹, Romaric Vincent¹, Fabrice Navarro², Mathilde Menneteau², Lamya Ghenim^3,4, Xavier Gidrol³, Thomas Bordy¹, Lionel Hervé¹, Olivier Cioni¹, Sabine Bardin⁵, Michel Bornens⁵, Yves Usson^4,6, Sophie Morales¹

¹CEA, LETI, DTBS, LISA, Université Grenoble Alpes, ²CEA, LETI, DTBS, LBAM, Université Grenoble Alpes, ³CEA, INSERM, BIG, Université Grenoble Alpes, ⁴CNRS, FR CNRS 3425, ⁵CNRS, UMR 144, Molecular Mechanisms of Intracellular Transport, PSL Research University, Institut Curie, ⁶TIMC-IMAG

Lens-free video microscopy enables us to monitor cell cultures directly inside the incubator. Here we describe the full protocol used to acquire and analyze a 2.7 day long acquisition of cultured HeLa cells, leading to a dataset of 2.2 x 10⁶ measurements of individual cell morphology and 10584 cell cycle tracks.

Biochemistry

Proteomic Analysis of Human Macrophage Polarization Under a Low Oxygen Environment

Magali Court^1,2, Marie Malier^1,2, Arnaud Millet^1,2,3

¹Team Mechanobiology, Immunity and Cancer, Institute for Advanced Biosciences, INSERM U1209, CNRS UMR5309, ²Université Grenoble Alpes, ³Pôle Recherche, Centre Hospitalier Universitaire des Alpes

We present a protocol to obtain proteomic signatures of human macrophages and apply this to determination of the impact of a low oxygen environment on macrophage polarization.

Biochemistry

Cell Culture on Silicon Nitride Membranes and Cryopreparation for Synchrotron X-ray Fluorescence Nano-analysis

Caroline Bissardon¹, Solveig Reymond¹, Murielle Salomé², Lionel André², Sam Bayat¹, Peter Cloetens², 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.

Chemistry

Biological Samples Preparation for Speciation at Cryogenic Temperature using High-Resolution X-Ray Absorption Spectroscopy

Caroline Bissardon¹, Marie-Pierre Isaure², Emmanuel Lesuisse³, 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.

Biology

Optimization of Crystal Growth for Neutron Macromolecular Crystallography

Elham Vahdatahar¹, Niels Junius^1,2, Monika Budayova - Spano¹

¹CEA, CNRS, IBS, Université Grenoble Alpes, ²ELVESYS SAS

Structural studies of biomacromolecules by crystallography require high-quality crystals. Here we demonstrate a protocol that can be used by OptiCrys (a fully automated instrument developed in our lab) and/or microdialysis buttons for growing large high-quality crystals based on knowledge of the crystallization phase diagram.

Biochemistry

High-Resolution Neutron Spectroscopy to Study Picosecond-Nanosecond Dynamics of Proteins and Hydration Water

Kevin Pounot^1,2, Markus Appel², Christian Beck^1,2, Martin Weik³, Giorgio Schirò³, Yann Fichou⁴, Tilo Seydel², Frank Schreiber¹

¹Institut für Angewandte Physik, Universität Tübingen, ²Institut Max von Laue - Paul Langevin (ILL), ³Institut de Biologie Structurale, Université Grenoble Alpes, ⁴Institut Européen de Chimie et Biologie, Bordeaux INP, Chimie et Biologie des Membranes et des Nanoobjets (CBMN), Université de Bordeaux

Neutron backscattering spectroscopy offers a nondestructive and label-free access to the ps-ns dynamics of proteins and their hydration water. The workflow is presented with two studies on amyloid proteins: on the time-resolved dynamics of lysozyme during aggregation and on the hydration water dynamics of tau upon fiber formation.