News Imagerie cellulaire - Cellular imaging
1.9K views | +3 today
Follow
Your new post is loading...
Scooped by RIC Paris-Saclay
Scoop.it!

Institut de Biologie Intégrative de la Cellule - Imagerie-Gif organizes FLIM-@-GIF (22 May 2PM)

Institut de Biologie Intégrative de la Cellule - Imagerie-Gif organizes FLIM-@-GIF (22 May 2PM) | News Imagerie cellulaire - Cellular imaging | Scoop.it

 

Imagerie-Gif Light Microscopy Facility

 

---------------------------------------------------------------------------------------------------

 

Wednesday 22th of May (14h00-17h00) listen to three experts to discover FLIM:


What is Fluorescence Lifetime Imaging Microscopy ?

            Fabienne Mérola (Laboratoire de Chimie Physique, Orsay)

 

Why using Fluorescence Lifetime Imaging in Biology ?

           Sergi Padilla-Parra (Medical Sciences Division, Univ of Oxford)

 

How to perform easy FLIM experiments?

           Clément Laigle (Leica-Microsystems)

 

Inscription is free but mandatory: Inscription link is here!

 

 ----------------------------------------------------------------------------------------------

Thursday 23th and Friday 24th of May: Workshops!


Imagerie-Gif will host for two days the Leica’s SP8 FALCON, an integrated solution for Fluorescence Lifetime Imaging that allows you to:

 

- Follow fast molecular interactions via FLIM-FRET

 

- Use biosensors to detect changes in metabolic state and

   microenvironment

 

- Apply lifetime contrast to separate multiple fluorophores

 

- Acquire fluorescence lifetime data with minimal training

 

Contact us to book a demonstration of the FALCON

and bring your own samples

 

-------------------------------------------------------------------

 

Contact: imagerie@i2bc.paris-saclay.fr

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Practical Course : Expansion Microscopy - 18/19 novembre - Gustave Roussy

Practical Course : Expansion Microscopy - 18/19 novembre - Gustave Roussy | News Imagerie cellulaire - Cellular imaging | Scoop.it

"La microscopie par dilatation"

 

 Cours pratique sur une technique de microscopie récente 

 

"L' Expansion Microsocopy" , c'est quoi ?  : 

 

Applicable sur tout type d’échantillon biologique, cette technique permet l’imagerie de détails très fins en les rendant physiquement plus grands grâce à un processus d'expansion chimique isotrope qui préserve les détails à l'échelle nanométrique (info :  Boyden E.S. 2015 Science)

 

Date : 18 et 19 Novembre

Lieu : Gustave Roussy

Inscription : gratuite mais limitée à 20 personnes

Dead line : 31 mai 2019

 Info et programme : site web de l’évènement.

 Contact : Tudor.MANOLIU - arobase - gustaveroussy.fr

 



more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

SAVE THE DATE   --  14 novembre 2019 - Journée RIC Paris-Saclay

SAVE THE DATE   --  14 novembre 2019 - Journée RIC Paris-Saclay | News Imagerie cellulaire - Cellular imaging | Scoop.it

Jeudi 14 novembre prochain aura lieu

la 5ème journée d’imagerie du RIC Paris-Saclay.

 

Trois thèmes : Imagerie ionique - Imagerie de la transcription - Imagerie de la mitochondrie

 

Lieu : Faculté de médecine Paris-Sud 

 

Programme : communiqué très prochainement.

 

Inscription : gratuite mais obligatoire

  

 

SAVE THE DATE : 14 NOVEMBRE 2019 -  (9h00 / 16h30)

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Supercritical Angle Fluorescence for Enhanced Axial Sectioning in STED Microscopy - ScienceDirect

Supercritical Angle Fluorescence for Enhanced Axial Sectioning in STED Microscopy - ScienceDirect | News Imagerie cellulaire - Cellular imaging | Scoop.it

 

Publication from UPSaclay

 

Sivankutty S, Hernández IC, Bourg N, Dupuis G, Lévêque-Fort S.

 

Institut des Sciences Moléculaires d’Orsay, CNRS, Univ. Paris-Sud, Université Paris-Saclay, F-91405 Orsay, France

Centre Laser de l’Université Paris Sud (CLUPS/LUMAT), Univ. Paris-Sud, CNRS, Université Paris-Saclay, F-91405 Orsay, France

 

We demonstrate subwavelength axial sectioning on biological samples with a stimulated emission depletion (STED) microscope combined with supercritical angle fluorescence (SAF) detection. SAF imaging is a powerful technique for imaging the membrane of the cell based on the direct exploitation of the fluorophore emission properties. Indeed, only when fluorophores are close to the interface can their evanescent near-field emission become propagative and be detected beyond the critical angle. Therefore, filtering out the SAF emission from the undercritical angle fluorescence (UAF) emission in the back focal plane of a high-NA objective lens permits nanometer axial sectioning of fluorescent emitters close to the coverslip. When combined with STED microscopy, a straightforward gain in axial resolution can be reached without any alteration of the STED beam path. Indeed, STED-SAF implementation only requires a modification in the detection path of the STED microscope and thus could be widely implemented.

 

Methods 2019  pii: S1046-2023(18)30427-4

https://doi.org/10.1016/j.ymeth.2019.03.027

PMID: 30946895.

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Multicolour single-molecule tracking of mRNA interactions with RNP granules

Multicolour single-molecule tracking of mRNA interactions with RNP granules | News Imagerie cellulaire - Cellular imaging | Scoop.it

Moon SL, Morisaki T, Khong A, Lyon K, Parker R, Stasevich TJ.

 

Ribonucleoprotein (RNP) granules are non-membrane-bound organelles that have critical roles in the stress response1,2, maternal messenger RNA storage3, synaptic plasticity4, tumour progression5,6 and neurodegeneration7,8,9. However, the dynamics of their mRNA components within and near the granule surface remain poorly characterized, particularly in the context and timing of mRNAs exiting translation. Herein, we used multicolour single-molecule tracking to quantify the precise timing and kinetics of single mRNAs as they exit translation and enter RNP granules during stress. We observed single mRNAs interacting with stress granules and P-bodies, with mRNAs moving bidirectionally between them. Although translating mRNAs only interact with RNP granules dynamically, non-translating mRNAs can form stable, and sometimes rigid, associations with RNP granules with stability increasing with both mRNA length and granule size. Live and fixed cell imaging demonstrated that mRNAs can extend beyond the protein surface of a stress granule, which may facilitate interactions between RNP granules. Thus, the recruitment of mRNPs to RNP granules involves dynamic, stable and extended interactions affected by translation status, mRNA length and granule size that collectively regulate RNP granule dynamics.

 

Nat Cell Biol. 2019; 21(2):162-168

Doi : https://doi.org/10.1038/s41556-018-0263-4

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Automating multimodal microscopy with NanoJ-Fluidics

Automating multimodal microscopy with NanoJ-Fluidics | News Imagerie cellulaire - Cellular imaging | Scoop.it

 

Pedro Almada, Pedro M. Pereira, Siân Culley, Ghislaine Caillol, Fanny Boroni-Rueda, Christina L. Dix, Guillaume Charras, Buzz Baum, Romain F. Laine, Christophe Leterrier & Ricardo Henriques


Combining and multiplexing microscopy approaches is crucial to understand cellular events, but requires elaborate workflows. Here, we present a robust, open-source approach for treating, labelling and imaging live or fixed cells in automated sequences. NanoJ-Fluidics is based on low-cost Lego hardware controlled by ImageJ-based software, making high-content, multimodal imaging easy to implement on any microscope with high reproducibility. We demonstrate its capacity on event-driven, super-resolved live-to-fixed and multiplexed STORM/DNA-PAINT experiments.

 

 

Nat Commun. 2019 Mar 15;10(1):1223

Open access : https://doi.org/10.1038/s41467-019-0923

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Correlative infrared nanospectroscopy and transmission electron microscopy to investigate nanometric amyloid fibrils: prospects and challenges.

Correlative infrared nanospectroscopy and transmission electron microscopy to investigate nanometric amyloid fibrils: prospects and challenges. | News Imagerie cellulaire - Cellular imaging | Scoop.it

publication from Paris-Saclay University

 

Partouche D, Mathurin J, Malabirade A, Marco S, Sandt C, Arluison V, Deniset-Besseau A, Trépout S.

 

1 Synchrotron SOLEIL, L'Orme des Merisiers Saint Aubin, Gif-sur-Yvette

2 Laboratoire Léon Brillouin LLB, CEA, CNRS UMR12, Université Paris Saclay

3 Laboratoire de Chimie Physique, CNRS, Univ. Paris-Sud, Université Paris-Saclay

4 INSERM, U1196, Université Paris Sud, Université Paris-Saclay

5 Institut Curie, PSL Research University, CNRS, UMR 9187

6 Université Paris Diderot-Paris 7, Sorbonne Paris Cité

 

Lay Description

Propagation of structural information through conformational changes in amyloid proteins is at the root of many neurodegenerative disorders. Amyloids are nanostructures originating from the aggregation of multiple copies of peptide or protein monomers that eventually form fibrils. Often described as being the cause for the development of various diseases, amyloid fibrils are of major significance in the public health domain. While important breakthroughs have been made in the field, fundamental issues like the 3D‐structures of the fibrils implied in some of those disorders are still to be elucidated. To better characterise these fibrils, a broad range of techniques is currently available for the detection and visualisation of amyloid nanostructures. Nevertheless none of them is able to perform direct chemical characterisation of single protein fibrils. In this work, we propose to investigate the structure of model amyloidogenic fibrils using a correlative approach. The complementary techniques used are transmission electron microscopy and a newly developed infrared nanospectroscopy technique called AFM‐IR that allows chemical characterisation at the nanometric scale. The strategy, protocol, challenges and difficulties encountered in this approach are introduced and discussed herein.

 

J Microsc. 2019 Jan 16

PMID : 30649833

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Recapitulating endocrine cell clustering in culture promotes maturation of human stem-cell-derived β cells

Recapitulating endocrine cell clustering in culture promotes maturation of human stem-cell-derived β cells | News Imagerie cellulaire - Cellular imaging | Scoop.it

Nair GG, Liu JS, Russ HA, Tran S, Saxton MS, Chen R, Juang C, Li ML, Nguyen VQ, Giacometti S, Puri S, Xing Y, Wang Y, Szot GL, Oberholzer J, Bhushan A and Hebrok M.

 

Diabetes Center, University of California San Francisco, San Francisco, CA, USA.

 

Despite advances in the differentiation of insulin-producing cells from human embryonic stem cells, the generation of mature functional β cells in vitro has remained elusive. To accomplish this goal, we have developed cell culture conditions to closely mimic events occurring during pancreatic islet organogenesis and β cell maturation. In particular, we have focused on recapitulating endocrine cell clustering by isolating and reaggregating immature β-like cells to form islet-sized enriched β-clusters (eBCs). eBCs display physiological properties analogous to primary human β cells, including robust dynamic insulin secretion, increased calcium signalling in response to secretagogues, and improved mitochondrial energization. Notably, endocrine cell clustering induces metabolic maturation by driving mitochondrial oxidative respiration, a process central to stimulus–secretion coupling in mature β cells. eBCs display glucose-stimulated insulin secretion as early as three days after transplantation in mice. In summary, replicating aspects of endocrine cell clustering permits the generation of stem-cell-derived β cells that resemble their endogenous counterparts.

 

Nat Cell Biol. 2019 Feb;21(2):263-274.

doi: 10.1038/s41556-018-0271-4

PubMed PMID: 30710150.

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

  A new 3D model of the NADPH oxidase cytosolic complex

   A new 3D model of the  NADPH  oxidase cytosolic complex | News Imagerie cellulaire - Cellular imaging | Scoop.it

From UPSaclay Laboratories

 

Cornelia S Ziegler, Leila Bouchab, Marc Tramier, Dominique Durand, Franck Fieschi, Sophie Dupré-Crochet, Fabienne Mérola, Oliver Nüße, Marie Erard

 

Laboratoire de Chimie Physique, CNRS, Univ. Paris-Sud, Université Paris-Saclay

Univ Rennes, CNRS, IGDR (Institut de génétique et développement de Rennes)

I2BC, CNRS, France

Universite Grenoble Alpes, IB

CNRS

Laboratoire de Chimie Physique, CNRS, Univ. Paris-Sud, Université Paris-Saclay

 

 

Phagocyte NADPH oxidase produces superoxide anions, a precursor of reactive oxygen species (ROS) critical for host responses to microbial infections. However, uncontrolled ROS production contributes to inflammation, making NADPH oxidase a major drug target. It consists of two membranous (Nox2 and p22phox) and three cytosolic subunits (p40phox, p47phox, and p67phox) that undergo structural changes during enzyme activation. Unraveling the interactions between these subunits and the resulting conformation of the complex could shed light on NADPH oxidase regulation and help identify inhibition sites. However, the structures and the interactions of flexible proteins comprising several well-structured domains connected by intrinsically disordered protein segments are difficult to investigate by conventional techniques such as X-ray crystallography, NMR, or cryo-EM. Here, we developed an analytical strategy based on FRET–fluorescence lifetime imaging (FLIM) and fluorescence cross-correlation spectroscopy (FCCS) to structurally and quantitatively characterize NADPH oxidase in live cells. We characterized the inter- and intramolecular interactions of its cytosolic subunits by elucidating their conformation, stoichiometry, interacting fraction, and affinities in live cells. Our results revealed that the three subunits have a 1:1:1 stoichiometry and that nearly 100% of them are present in complexes in living cells. Furthermore, combining FRET data with small-angle X-ray scattering (SAXS) models and published crystal structures of isolated domains and subunits, we built a 3D model of the entire cytosolic complex. The model disclosed an elongated complex containing a flexible hinge separating two domains ideally positioned at one end of the complex and critical for oxidase activation and interactions with membrane components.

 

J Biol Chem. 2019 Jan 10. pii: jbc.RA118.006864

doi: 10.1074/jbc.RA118.006864.

Open Access : www.jbc.org/content/early/2019/01/10/jbc.RA118.006864.full.pdf?with-ds=yes

 

 

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

EB1 contributes to microtubule bundling and organization, along with root growth, in Arabidopsis thaliana

EB1 contributes to microtubule bundling and organization, along with root growth, in Arabidopsis thaliana | News Imagerie cellulaire - Cellular imaging | Scoop.it
From the Université Paris-Saclay
 
Arthur T. Molines1, Jessica Marion1, Salem Chabout2, Laetitia Besse3, Jim P. Dompierre3, Grégory Mouille2, Frédéric M. Coquelle1
 

1  Department of Cell Biology, Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France

2  Institut Jean-Pierre Bourgin (IJPB), INRA - AgroParisTech, 78026 Versailles Cedex, France

3  Light Microscopy Facility, Imagerie-Gif, Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France

 

 

 

Microtubules are involved in plant development and adaptation to their environment, but the sustaining molecular mechanisms remain elusive. Microtubule-end-binding 1 (EB1) proteins participate in directional root growth in Arabidopsis thaliana . However, a connection to the underlying microtubule array has not been established yet. We show here that EB1 proteins contribute to the organization of cortical microtubules in growing epidermal plant cells, without significant modulation of microtubule dynamics. Using super-resolution stimulated emission depletion (STED) microscopy and an original quantification approach, we also demonstrate a significant reduction of apparent microtubule bundling in cytoplasmic-EB1-deficient plants, suggesting a function for EB1 in the interaction between adjacent microtubules. Furthermore, we observed root growth defects in EB1-deficient plants, which are not related to cell division impairment. Altogether, our results support a role for EB1 proteins in root development, in part by maintaining the organization of cortical microtubules.

 

Biol Open. 2018 Aug 2;7(8)

DOI : https://doi.org/10.1242/bio.030510

PubMed : 29945874

Open access : http://bio.biologists.org/content/biolopen/7/8/bio030510.full.pdf

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Optoacoustic microscopy at multiple discrete frequencies

Optoacoustic microscopy at multiple discrete frequencies | News Imagerie cellulaire - Cellular imaging | Scoop.it

Stephan Kellnberger, Dominik Soliman, George J. Tserevelakis, Markus Seeger, Hong Yang, Angelos Karlas, Ludwig Prade, Murad Omar & Vasilis Ntziachristos

 

Optoacoustic (photoacoustic) sensing employs illumination of transient energy and is typically implemented in the time domain using nanosecond photon pulses. However, the generation of high-energy short photon pulses requires complex laser technology that imposes a low pulse repetition frequency (PRF) and limits the number of wavelengths that are concurrently available for spectral imaging. To avoid the limitations of working in the time domain, we have developed frequency-domain optoacoustic microscopy (FDOM), in which light intensity is modulated at multiple discrete frequencies. We integrated FDOM into a hybrid system with multiphoton microscopy, and we examine the relationship between image formation and modulation frequency, showcase high-fidelity images with increasing numbers of modulation frequencies from phantoms and in vivo, and identify a redundancy in optoacoustic measurements performed at multiple frequencies. We demonstrate that due to high repetition rates, FDOM achieves signal-to-noise ratios similar to those obtained by time-domain methods, using commonly available laser diodes. Moreover, we experimentally confirm various advantages of the frequency-domain implementation at discrete modulation frequencies, including concurrent illumination at two wavelengths that are carried out at different modulation frequencies as well as flow measurements in microfluidic chips and in vivo based on the optoacoustic Doppler effect. Furthermore, we discuss how FDOM redefines possibilities for optoacoustic imaging by capitalizing on the advantages of working in the frequency domain.

 

Light: Science & Applications volume 7, Article number: 109 (2018)

https://doi.org/10.1038/s41377-018-0101-2

Open Access : https://www.nature.com/articles/s41377-018-0101-2.pdf

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Deep learning enables cross-modality super-resolution in fluorescence microscopy

Deep learning enables cross-modality super-resolution in fluorescence microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it

Hongda Wang, Yair Rivenson, Yiyin Jin, Zhensong Wei, Ronald Gao, Harun Günaydın, Laurent A. Bentolila, Comert Kural & Aydogan Ozcan

 

We present deep-learning-enabled super-resolution across different fluorescence microscopy modalities. This data-driven approach does not require numerical modeling of the imaging process or the estimation of a point-spread-function, and is based on training a generative adversarial network (GAN) to transform diffraction-limited input images into super-resolved ones. Using this framework, we improve the resolution of wide-field images acquired with low-numerical-aperture objectives, matching the resolution that is acquired using high-numerical-aperture objectives. We also demonstrate cross-modality super-resolution, transforming confocal microscopy images to match the resolution acquired with a stimulated emission depletion (STED) microscope. We further demonstrate that total internal reflection fluorescence (TIRF) microscopy images of subcellular structures within cells and tissues can be transformed to match the results obtained with a TIRF-based structured illumination microscope. The deep network rapidly outputs these super-resolved images, without any iterations or parameter search, and could serve to democratize super-resolution imaging.

 

Nature Methods (2018) Published: 17 December 2018

https://doi.org/10.1038/s41592-018-0239-0

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

CNRS on Instagram: “#RETRO2017 Replis de la #muqueuse intestinale d’une souris observés en #microscopie confocale multi-couleurs. ”

CNRS on Instagram: “#RETRO2017 Replis de la #muqueuse intestinale d’une souris observés en #microscopie confocale multi-couleurs. ” | News Imagerie cellulaire - Cellular imaging | Scoop.it
more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Insight into microtubule nucleation from tubulin-capping proteins

Insight into microtubule nucleation from tubulin-capping proteins | News Imagerie cellulaire - Cellular imaging | Scoop.it

Publication from Paris-Saclay


Valérie Campanacci, Agathe Urvoas, Soraya Cantos-Fernandes, Magali Aumont-Nicaise, Ana-Andreea Arteni, Christophe Velours, Marie Valerio-Lepiniec, Birgit Dreier, Andreas Plückthun, Antoine Pilon, Christian Poüs, Philippe Minard, and Benoît Gigant

 

Institute for Integrative Biology of the Cell , CEA, CNRS,  Univ Paris-Saclay, Gif-sur-Yvette 
Department of Biochemistry, University of Zurich, CH-8057 Zurich, Switzerland
INSERM UMR-S 1193, Université Paris-Sud, Université Paris-Saclay, Châtenay-Malabry
Biochimie, APHP, Hôpitaux Universitaires de l'Est Parisien
INSERM UMR-S 1193, Université Paris-Saclay, Châtenay-Malabry

Biochimie-Hormonologie, APHP, Hôpitaux Universitaires Paris-Sud, Clamart

  

correspondence  : christian.pous@u-psud.fr   --   benoit.gigant@i2bc.paris-saclay.fr.

 


Significance
Microtubules are involved in many key functions of eukaryotic cells, including cell division, intracellular transport, and cell shape. They are hollow tubes made of parallel filaments, themselves formed by the self-assembly of αβ-tubulin molecules. Whereas microtubules lengthen and shorten from their ends dynamically, their birth, called nucleation, remains poorly understood. To gain information on this process, we have determined the structure of tubulin bound to CopN, a bacterial protein that delays nucleation. Together with the behavior of artificial tubulin-binding proteins, our results lead to the hypothesis that targeting two filaments at the fast-growing end of the microtubule inhibits nucleation. They also suggest different dynamics at both ends of the nucleus.

 

 

Proc Natl Acad Sci U S A. 2019 Apr 29. pii: 201813559.

https://doi.org/10.1073/pnas.1813559116

PMID :31036638

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Combining 3D single molecule localization strategies for reproducible bioimaging

Combining 3D single molecule localization strategies for reproducible bioimaging | News Imagerie cellulaire - Cellular imaging | Scoop.it

Publication from Paris-Saclay

 

Clément Cabriel, Nicolas Bourg, Pierre Jouchet, Guillaume Dupuis, Christophe Leterrier, Aurélie Baron, Marie-Ange Badet-Denisot, Boris Vauzeilles, Emmanuel Fort & Sandrine Lévêque-Fort

 

Institut des Sciences Moléculaires d'Orsay, CNRS,Université Paris-Saclay

Centre de Photonique BioMédicale, Université Paris-Saclay, CNRS, Fédération LUMAT

Aix-Marseille Université, CNRS, INP, NeuroCyto, Marseille

Centre de Recherche de Gif, Institut de Chimie des Substances Naturelles du CNRS, Gif-sur-Yvette

Laboratoire de Synthèse de Biomolécules, Institut de Chimie Moléculaire et des Matériaux d'Orsay, Université Paris-Saclay, CNRS.

Institut Langevin, ESPCI Paris, PSL University, CNRS

Institut des Sciences Moléculaires d'Orsay, CNRS, Université Paris-Saclay


Here, we present a 3D localization-based super-resolution technique providing a slowly varying localization precision over a 1 μm range with precisions down to 15 nm. The axial localization is performed through a combination of point spread function (PSF) shaping and supercritical angle fluorescence (SAF), which yields absolute axial information. Using a dual-view scheme, the axial detection is decoupled from the lateral detection and optimized independently to provide a weakly anisotropic 3D resolution over the imaging range. This method can be readily implemented on most homemade PSF shaping setups and provides drift-free, tilt-insensitive and achromatic results. Its insensitivity to these unavoidable experimental biases is especially adapted for multicolor 3D super-resolution microscopy, as we demonstrate by imaging cell cytoskeleton, living bacteria membranes and axon periodic submembrane scaffolds. We further illustrate the interest of the technique for biological multicolor imaging over a several-μm range by direct merging of multiple acquisitions at different depths.

 

Nat Commun. 2019, 30;10(1):1980

https://doi.org/10.1038/s41467-019-09901-8

PMID: 31040275

Open Acces : https://www-nature-com.gate2.inist.fr/articles/s41467-019-09901-8.pdf

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

3D + time imaging of normal and twin sea urchin embryos for the reconstruction of their cell lineage 

3D + time imaging of normal and twin sea urchin embryos for the reconstruction of their cell lineage  | News Imagerie cellulaire - Cellular imaging | Scoop.it

 

Publication from UPSaclay

 

Antonio Ortiz, Elena Kardash and Nadine Peyrieras

 

BioEmergences Laboratory (USR3695), CNRS, Université Paris-Saclay, Gif-sur-Yvette, France

 

The Mediterranean sea urchin, Paracentrotus lividus, has been a powerful model to study embryonic development since the late 1800s. As a model, it has the advantage of having external fertilization, it can easily be manipulated experimentally, and it has semi-transparent embryonic stages, which makes it ideal for live imaging. Embryogenesis is a highly dynamic process with intrinsic variability. The reconstruction of cell dynamics and an assessment of such variability from in vivo observations has proven to be a challenge. Here, we provide an innovative methodology for manipulation and immobilization of embryos and their long-term 3D + time imaging. We then describe the twinning procedure that allows us to assess the variability and robustness of developmental processes. We demonstrate the reconstruction of cell lineages based on automated image processing and cell tracking using the BioEmergences workflow as well as the use of interactive visualization tools (Mov-IT software) for lineage validation, correction and analysis.

 

 

Methods Cell Biol. 2019;151:399-418

https://doi.org/10.1016/bs.mcb.2019.01.008

PMID: 30948021.

 

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

About samples, giving examples: Optimized Single Molecule Localization Microscopy

About samples, giving examples: Optimized Single Molecule Localization Microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it

Angélique Jimenez, Karoline Friedl, Christophe Leterrier

 

 

Super-resolution microscopy has profoundly transformed how we study the architecture of cells, revealing unknown structures and refining our view of cellular assemblies. Among the various techniques, the resolution of Single Molecule Localization Microscopy can reach the size of macromolecular complexes and offer key insights on their nanoscale arrangement in situ. SMLM is thus a demanding technique and taking advantage of its full potential requires specifically optimized procedures. Here we describe how we perform the successive steps of an SMLM workflow, focusing on single-color Stochastic Optical Reconstruction Microscopy (STORM) as well as multicolor DNA Points Accumulation for imaging in Nanoscale Topography (DNA-PAINT) of fixed samples. We provide detailed procedures for careful sample fixation and immunostaining of typical cellular structures: cytoskeleton, clathrin-coated pits, and organelles. We then offer guidelines for optimal imaging and processing of SMLM data in order to optimize reconstruction quality and avoid the generation of artifacts. We hope that the tips and tricks we discovered over the years and detail here will be useful for researchers looking to make the best possible SMLM images, a pre-requisite for meaningful biological discovery.

 

DOI : http://dx.doi.org/10.1101/568295

Open access :https://www.biorxiv.org/content/biorxiv/early/2019/03/05/568295.full-text.pdf

 

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Coming together to define membrane contact sites

Coming together to define membrane contact sites | News Imagerie cellulaire - Cellular imaging | Scoop.it

Scorrano L, De Matteis MA, Emr S, Giordano F, Hajnóczky G, Kornmann B, Lackner LL, Levine TP, Pellegrini L, Reinisch K, Rizzuto R, Simmen T, Stenmark H, Ungermann C, Schuldiner M.

 

 

Given the recent growing interest in interorganelle membrane contact sites, the field will benefit from clear rules to define and study them. In this Perspective, a panel of experts aims to provide this growing field with guidelines for experimental definition and analysis.

 

 

 

 

Nat Commun. 2019, 10(1):1287

doi: https://doi.org/10.1038/s41467-019-09253-3

Open acces : https://www.nature.com/articles/s41467-019-09253-3.pdf

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Human cytomegalovirus hijacks the autophagic machinery and LC3 homologs in order to optimize cytoplasmic envelopment of mature infectious particles

Human cytomegalovirus hijacks the autophagic machinery and LC3 homologs in order to optimize cytoplasmic envelopment of mature infectious particles | News Imagerie cellulaire - Cellular imaging | Scoop.it

publication from UPSaclay

 

Clémence Taisne, Marion Lussignol, Eva Hernandez, Arnaud Moris, Lina Mouna & Audrey Esclatine

 

1. Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, Gif-sur-Yvette , France.

2. Sorbonne Université, Centre d'Immunologie et des Maladies Infectieuses, INSERM U1135, CNRS ERL 8255, Paris, France.

3. Virologie, APHP, Hôpital Paul Brousse, 94800, Villejuif, France.

4. Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette cedex, France.



During its life cycle, Human cytomegalovirus (HCMV) tightly modulates autophagy, a vesicular pathway allowing degradation and recycling of cellular components. To study the interplay between autophagy and the viral life cycle, we established various autophagy-deficient human fibroblastic cell lines. By knocking down the expression or activity of five autophagy-related proteins, we confirmed the proviral function that the autophagic machinery exerts on HCMV production. Using 3D reconstruction from confocal microscopy and electron microscopy, we demonstrated that lipidated LC3-positive vesicles accumulated at the viral assembly compartment (vAC). The vAC is a juxtanuclear ring-shaped structure containing several organelles and membranes, where assembly and final envelopment of HCMV particles occur. Two LC3 homologs, GABARAPL1 and GATE16, also accumulated during HCMV infection and were associated with the vAC, in proximity with fragmented Golgi stacks. Additionally, we observed the formation of a pre-assembly compartment (PrAC) in infected cells, which consists of a juxtanuclear structure containing both fragmented Golgi and LC3-positive vesicles. Finally, we showed that highly purified extracellular viral particles were associated with various autophagy proteins. Our results thus suggest that autophagy machinery participates to the final cytoplasmic envelopment of HCMV viral particles into the vAC and that autophagy-related proteins can be spotted in the virions.

 

Sci Rep. 2019, 9(1):4560

Open acces : https://doi.org/10.1038/s41598-019-41029-z

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Early mitochondrial fragmentation is a potential in vitro biomarker of environmental stress.

Early mitochondrial fragmentation is a potential in vitro biomarker of environmental stress. | News Imagerie cellulaire - Cellular imaging | Scoop.it

publication from UPSaclay

  

Perdiz Daniel 1, Oziol Lucie 2, Poüs Christian 3

 

1 Univ. Paris-Sud, INSERM UMR-S 1193, Université Paris Sud-Paris Saclay, Faculté de Pharmacie, Châtenay-Malabry, France

2 Univ. Paris-Sud, CNRS UMR 8079, Université Paris Sud-Paris Saclay, Faculté de Pharmacie, Châtenay-Malabry, France

3 Biochimie-Hormonologie, APHP, Hôpitaux Universitaires Paris-Sud, Site Antoine Béclère, Clamart, France

 

 

Mitochondria are essential dynamic organelles that ordinarily balance between fragmentation and fusion. Under stress conditions, a shift toward fragmentation or hyper-fusion is observed as a pro-survival reaction. Fragmentation of mitochondria occurs within minutes or hours after the beginning of the stress and occurs in response to a large number of stress stimuli, including those triggered by environmental contaminants. In this study, we tested whether the change in the mitochondrial phenotype, from tubular to fragmented, could be used as a potential environmental stress biomarker in cells and compared this response with the standard MTT-based viability assay. Firstly, we show that mitochondrial fragmentation induced by selected stressors not only increases with concentrations, but also correlates positively with the cytotoxicity. Secondly, we found that the mitochondrial fragmentation that occurs in the first hour of stress correlated with the viability measured after a 24-h stress, allowing the establishment of a linear relation between mitochondrial fragmentation at 1 h and the predictable associated cytotoxicity of environmental contaminants alone or in mixture. In conclusion, we have succeeded in developing a model of predictable 24 h-cytotoxicity given mitochondrial fragmentation at 1 h with a set of chemicals. This model has been successful applied to three environmental toxicants and to a set of two chemical mixtures. We thus propose that mitochondrial fragmentation is a response that could be used as an early in vitro biomarker of environmental stress for toxicants alone or in mixture.

 

 

Chemosphere. 2019 Feb 15;223:577-587

https://doi.org/10.1016/j.chemosphere.2019.02.044

PubMed PMID: 30797167.

 

 

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Événements | Université Paris Saclay

Événements | Université Paris Saclay | News Imagerie cellulaire - Cellular imaging | Scoop.it

Université Paris-Saclay events

 

Paris-Saclay | Meet my Platform : video

 

 

Événements

 

Une journée de rencontres plateformes entreprises organisée par Genopole et le Département Sciences de la Vie de l'Université Paris-Saclay
 

Le 31 janvier 2019,  Meet My Platform | Université Paris-Saclay, les Sciences de la Vie vous font découvrir leurs plateformes.

Vous faites partie d'une plateforme Sciences de la Vie de l'Université Paris-Saclay et vous souhaitez découvrir les autres plateformes du territoire ? Vous souhaitez vous faire connaître auprès d'elles et des entreprises ? Venez présenter vos savoir-faire et équipements lors de cette journée.

 

Vous êtes une entreprise, une unité de recherche et vous avez besoin d'utiliser des équipements de pointe en Sciences de la Vie ? Vous souhaitez élargir votre réseau de partenaires académiques dans ce domaine ? Venez découvrir le potentiel des plateformes technologiques de l'Université Paris-Saclay.

Programme :

9h30 : accueil

 

10h00 : ouverture de l'événement par Genopole et l'Université Paris-Saclay

 

10h20 – 11h00 : table ronde "la vision de trois industriels sur le potentiel des plateformes Sciences de la Vie de l'Université Paris-Saclay" avec :

  • Laurent CHENE, Head of drug discovery, ENTEROME

  • Georges DA VIOLANTE, Metabolism Dept Manager, in charge of academic and industrial partnerships, SERVIER

  • Artem KHLEBNIKOV, Directeur Stratégique Partnerships, DANONE

Table ronde animée par Nathalie Tourret, journaliste

 

11h00 – 12h00 : découverte de 4 réseaux / typologies sélectionnées de plateformes

 

12h00 : buffet

 

12h30 - 14h00 : stands et networking

 

14h00 – 17h00 : rendez-vous B2B

 

14h00 – 17h00 : ateliers pour les plateformes

 

Cette journée est organisée par Genopole et le Département Sciences de la Vie de l'Université Paris-Saclay, avec le soutien du pôle de compétitivité Medicen Paris Région, de la SATT Paris-Saclay et de CentraleSupélec

 

La matinée aura lieu dans l'auditorium Michelin bâtiment Eiffel et l'après-midi au rez-de-chaussée du bâtiment Bouygues de CentraleSupélec (les deux espaces sont l'un en face l'autre).

 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Quantification of protein mobility and associated reshuffling of cytoplasm during chemical fixation

Quantification of protein mobility and associated reshuffling of cytoplasm during chemical fixation | News Imagerie cellulaire - Cellular imaging | Scoop.it

Jan Huebinger, Jessica Spindler, Kristin J. Holl and  Björn Koos

 

Department of Systemic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Str.11, 44227, Dortmund, Germany.

 

To understand cellular functionalities, it is essential to unravel spatio-temporal patterns of molecular distributions and interactions within living cells. The technological progress in fluorescence microscopy now allows in principle to measure these patterns with sufficient spatial resolution. However, high resolution imaging comes with long acquisition times and high phototoxicity. Therefore, physiological live cell imaging is often unfeasible and chemical fixation is employed. Yet, fixation methods have not been rigorously investigated, in terms of pattern preservation, at the resolution at which cells can now be imaged. A key parameter for this is the time required until fixation is complete. During this time, cells are under unphysiological conditions and patterns decay. We demonstrate here that formaldehyde fixation takes more than one hour for cytosolic proteins in cultured cells. Other small aldehydes, glyoxal and acrolein, did not perform better. Associated with this, we found a distinct displacement of proteins and lipids, including their loss from cells. Fixations using glutaraldehyde were faster than four minutes and retained most cytoplasmic proteins. Surprisingly, autofluorescence produced by glutaraldehyde was almost completely absent with supplementary addition of formaldehyde without compromising fixation speed. These findings indicate, which cellular processes can actually be reliably imaged after a certain chemical fixation.

 

Sci Rep. 2018; 8: 17756

doi : 10.1038/s41598-018-36112-w

Open access article : https://www.nature.com/articles/s41598-018-36112-w.pdf

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Multifocal multiphoton volumetric imaging approach for high-speed time-resolved Förster resonance energy transfer imaging in vivo

Multifocal multiphoton volumetric imaging approach for high-speed time-resolved Förster resonance energy transfer imaging in vivo | News Imagerie cellulaire - Cellular imaging | Scoop.it

Simon P. Poland, Grace K. Chan, James A. Levitt, Nikola Krstajić, Ahmet T. Erdogan, Robert K. Henderson, Maddy Parsons, and Simon M. Ameer-Beg

 

In this Letter, we will discuss the development of a multifocal multiphoton fluorescent lifetime imaging system where four individual fluorescent intensity and lifetime planes are acquired simultaneously, allowing us to obtain volumetric data without the need for sequential scanning at different axial depths. Using a phase-only spatial light modulator (SLM) with an appropriate algorithm to generate a holographic pattern, we project a beamlet array within a sample volume of a size, which can be preprogrammed by the user. We demonstrate the capabilities of the system to image live-cell interactions. While only four planes are shown, this technique can be rescaled to a large number of focal planes, enabling full 3D acquisition and reconstruction.

 

Optics Letters Vol. 43, Issue 24, pp. 6057-6060 (2018)

https://doi.org/10.1364/OL.43.006057

Open Access : https://www.osapublishing.org/ol/viewmedia.cfm?uri=ol-43-24-6057&seq=0 

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Super-resolution enhancement by quantum image scanning microscopy

Super-resolution enhancement by quantum image scanning microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it

Ron Tenne, Uri Rossman, Batel Rephael, Yonatan Israel, Alexander Krupinski-Ptaszek, Radek Lapkiewicz, Yaron Silberberg & Dan Oron

 

The principles of quantum optics have yielded a plethora of ideas to surpass the classical limitations of sensitivity and resolution in optical microscopy. While some ideas have been applied in proof-of-principle experiments, imaging a biological sample has remained challenging, mainly due to the inherently weak signal measured and the fragility of quantum states of light. In principle, however, these quantum protocols can add new information without sacrificing the classical information and can therefore enhance the capabilities of existing super-resolution techniques. Image scanning microscopy, a recent addition to the family of super-resolution methods, generates a robust resolution enhancement without reducing the signal level. Here, we introduce quantum image scanning microscopy: combining image scanning microscopy with the measurement of quantum photon correlation allows increasing the resolution of image scanning microscopy up to twofold, four times beyond the diffraction limit. We introduce the Q-ISM principle and obtain super-resolved optical images of a biological sample stained with fluorescent quantum dots using photon antibunching, a quantum effect, as a resolution-enhancing contrast mechanism.

 

Nature Methods (2018) Published: 17 December 2018

https://doi.org/10.1038/s41566-018-0324-z

more...
No comment yet.
Scooped by RIC Paris-Saclay
Scoop.it!

Imaging cellular ultrastructures using expansion microscopy (U-ExM)

Imaging cellular ultrastructures using expansion microscopy (U-ExM) | News Imagerie cellulaire - Cellular imaging | Scoop.it

Davide Gambarotto, Fabian U. Zwettler, Maeva Le Guennec, Marketa Schmidt-Cernohorska, Denis Fortun, Susanne Borgers, Jörn Heine, Jan-Gero Schloetel, Matthias Reuss, Michael Unser, Edward S. Boyden, Markus Sauer, Virginie Hamel & Paul Guichard

 

Determining the structure and composition of macromolecular assemblies is a major challenge in biology. Here we describe ultrastructure expansion microscopy (U-ExM), an extension of expansion microscopy that allows the visualization of preserved ultrastructures by optical microscopy. This method allows for near-native expansion of diverse structures in vitro and in cells; when combined with super-resolution microscopy, it unveiled details of ultrastructural organization, such as centriolar chirality, that could otherwise be observed only by electron microscopy.

 

Nature Methods (2018) Published: 17 December 2018

https://doi.org/10.1038/s41592-018-0238-1

more...
No comment yet.