PIBBS-MARS & AFM comprises two facilities :

• The AFM facility is the only one in the FBI Infrastructure to provide access to state-of-the-art Atomic Force Microscopes, including a custom built high-speed AFM.
• The MARS facility is devoted to cutting-edge optical microscopies such as Single Molecule Localization Microscopy, smFRET, PIE-FCCS, 2-photons FCS, single particle tracking, etc… The MARS R&D division, closely associated to two R&D teams, is in charge of implementing and developing new custom
  advanced microscopies (such as STED-FCCS, 2foci-FCS or multifocal microscopy) before their transfer to the facility.

Moreover, the AFM and MARS facilities and staff work closely together to develop and transfer new correlative imaging modalities, such as AFM / Superresolution or AFM / Confocal (FLIM, spFRET).

Users are assisted by dedicated research engineers and scientific coordinators to define the best approach, experimental design, help in data acquisition and analysis.

http://www.cbs.cnrs.fr/index.php/fr/fluorescence

Microscopy systems available @PIBBS

The H2P2 platform performs histology work on human, animal and plant tissues. The facility is equipped with last generation devices to increase analysis reproducibility and ensure rapid automatic processing. We produce Tissues Micro Array (TMA) that can group hundreds of tissulars spots on a single microscope slide. We have developed a thorough expertise in immuno-labeling (more than 1000 Ac in our catalog) as well as multiplex staining, a technique that can highlight up to 6 different proteins per slide, fluorescence or chromogenic. Since 2021, with the CELLDive technology we are able to visualize up to 60 markers on the same sample. To visualize slides we offer several options: epifluorescence microscope, virtual slides from a Hamamatsu scanner with fluorescence and a new generation confocal scanner (3D-Histech) with seven channels in fluorescence.  This latter technology gives the opportunity to scan thick tissues (300µm) after clearing. Slides are analysed in 2D using Halo, an image analysis platform/software (with machine learning) for quantitative tissue analysis in digital pathology.  We perform 3D images routinely and advanced analysis reconstruction with Amira. We also developed extensive expertise in laser capture microdissection, allowing single cells capture. We are developing in situ molecular technologies, as Raman micro spectroscopy imaging on biological samples with statistical analysis of obtained spectra.

MicroPICell is an imaging facility specialized in sample preparation (histology), cellular and tissular imaging (photonic microscopy) and data processing and analysis (bioimage analysis). The facility is located within the precinct of the Hotel Dieu hospital, Nantes University. The platform is affiliated to the Structure Federative de Recherche UMS INSERM 016/CNRS 3556/Nantes University and teamed up with Nikon Instrument society and the APEX veterinary facility to create the Center of Excellence Nikon Nantes in 2016. The MicroPICell facility offers a complete integrated cellular imaging service to all users after analysis of the feasibility of their projects. The applications range is then very large, for example cancer research and immunology, regenerative medicine or stem cells research. It is organized in 3 main services: histology, photonic microscopy, image data management and analysis, all these services working closely together on users projects.

The Imaging and Cytometry Platform (PFIC) is one of the 9 scientific Platforms of the UMS AMMICa of Gustave Roussy, one of the first European comprehensive cancer center, located in the south of Paris.
Supporting basic and clinical research programs on cancer, the PFIC is a service, training and R&D center at the interface of basic, translational and clinical research.
The PFIC provides research and industry with an open center of expertise in multi-scale photonic imaging from molecular to tissue, and from animal models to the human. Run by 9 engineers from which 5 dedicated to imaging, the PFIC is organized into specialized units to offer expertise:

• In confocal and multiphoton imaging together with the combined techniques TIRF, live SR, FRAP, FRET, photoconversion, for the study of dynamic interactions at high resolution.
• In complex multidimensional dynamic imaging in living organs, 3D-organoid models, high resolution intravital imaging on small animal and whole animal imaging.
• In transfer of photonics into the clinic (New contrast, NIR and confocal)
• In flow, spectral and mass cytometry and high throughput cell sorting and cloning
• Bioinformatics expertise for data processing and quantification

The PFIC is also strongly involved, with industrial partners, in innovative developments in new optical devices, new fluorescent probes and specific requests for clinical transfer of photon imaging.

Microscopy systems available @PFIC

The Morphoscope / Polytechnique BioImaging Facility received the IBiSA labelled in 2019. It is hosted and operated by the Laboratory for Optics & Biosciences (LOB) at Ecole Polytechnique, one of the founding members of the France BioImaging infrastructure. The Morphoscope facility results from a major equipment investment grant initiated in 2013 (Equipex Morphoscope2 project). It is based on the expertise of the LOB Advanced Microscopies group (9 tenured scientists, 1 tenured research engineer, 1 zootechnician, students, and access to LOB electronics & mechanics workshops). One important specificity of this platform is its emphasis on technology development and methodological innovation. In particular, the host team is a world leader in multiphoton microscopy of tissues, and the platform hosts both state-of-the- art commercial systems and lab-built systems based on cutting-edge technologies.
Imaging equipment is organized along three complementary needs:

•  deep tissue imaging with multi-contrast nonlinear microscopy;
• fast imaging of semi-transparent samples using light-sheet excitation microscopy;
• super-resolution optical microscopy.

In addition, we have an expertise in large-volume bioimage informatics (e.g. managing & processing 3D images with >10^10 voxels)
We can provide the following:

• Case study, expertise, and technological innovation in cell and tissue imaging.
• Tissue imaging:
  • multicontrast multiphoton microscopy: 2PEF, SHG, THG, polarimetry, FLIM, CARS, multicolor 2-photon through wavelength mixing;
  • deep-tissue three-photon microscopy (Light Sci App 2018);
  • large volume multicolor microscopy of uncleared tissue ex vivo (ChroMS, multicolor 2P integrated with automated serial cutting, Nat Commun 2019);
• Fast imaging:
  • standard and 2-photon light-sheet microscopy (Nat Methods 2014).
• Super-resolved optical imaging:
  • TIRF-SIM microscopy.

Installed equipment:

Application-oriented setups (access through collaborations or reservation):

• Multimodal/multicolor multiphoton microscope (2PEF, SHG, THG) – LaVision Biotec, Trimscope-II equipped with three femtosecond beams.
• Super-resolution SIM / TIRF microscope – GE Healthcare, DeltaVision OMX SR.
• Confocal microscope (FRAP, white light laser, resonant scanner) – Leica, TCS SP 8X.
• Multi-view light-sheet microscope – Luxendo-Bruker, MuVi-SPIM.

Methodology-oriented setups (collaborative access):

• Multimodal multiphoton microscope (2PEF, SHG, THG, FLIM, CARS, polar.) – Lab-built.
• Multicolor 2PEF microscope (wavelength mixing, brainbow) – Lab-built.
• 3-photon/THG microscope – Lab-built.
• Two-photon light-sheet microscope – Lab-built.

Zebrafish facility. Capacity 6000 fish.
Secured image server with an OMERO installation (70 TB).
One GPU compute server with NVidia v100x3
Several image processing/analysis workstations, including 3 Imaris licences.

Microscopy systems available @Morphoscope

InMagiC (INMED iMAGing Center) is the microscopy platform of the Institute of Neurobiology of the Mediterranean: INMED in Marseille on the Luminy campus.
The platform is under the scientific direction of Rosa COSSART and under the responsibility of the engineer François MICHEL rosa.cossart@inserm.fr / francois.michel@inserm.fr

INMAGIC gathers in one place the multi-scale study tools necessary to study the development and the physio-pathological functioning of the central nervous system.
These tools cover the scales of observation from the whole animal (binocular loupes) to the dynamic morphology of neuronal contacts (synapses) thanks to widefield, confocal and multiphoton microscopes.
In addition, the platform provides a large number of training courses at all levels, both theoretical and practical, for all audiences (students, engineers, researchers, etc.).

The strong and original points of our structure are:

• A space dedicated to the techniques of tissue clearing and the acquisition of these samples by an ultramicroscope (light sheet) in platform access, as well as the expertise of the engineer in charge (active member of the GT transparencies of the RTMFM);
• The use of the multiphoton microscopy for the fast measurement (10-20 Hz) of the neuronal activity in vitro and in vivo on large populations of cells (up to more than 500 simultaneous cells).

Two high performance analysis stations are also available to all users. They include free and commercial software dedicated to analysis such as FIJI, ICY, IMARIS, Neurolucida360, Matlab…

Finally, the platform is a laureate of the PIA Equipex + 2020 and in this context the availability of tools beyond the state of the art in minimally invasive in vivo imaging will soon be open to the community.
These new tools will be commercial with industrial co-developments: a 3-photon excitation microscope, an ultra-fast AOD scanning microscope, a 2-photon endoscope or in academic development with the collaboration of the Fresnel Institute such as an ultra-thin endoscope and a deep recording station by photo-acoustics.

The ISO 9001 certified platform consists of 2 systems:

• Zeiss Axio Imager M2 LSM 800 confocal upright microscope with PI XY motorized piezo stage, 2 PMTs and an AiryScan super resolution module The excitation wavelengths available on the equipment are 405nm, 488nm, 561 nm, 640 nm. An OKOLAB incubation chamber maintains a stable temperature and CO2 level in the microscope for live image acquisition. This equipment allows the acquisition of topography information by reflection and fluorescence simultaneously.
• Inverted confocal microscope Spinning disk CSU W1: The stand is an Eclipse Ti2-E Nikon equipped with a motorized XY stage and a piezzo Z-stage, the “Perfect Focus” focus maintenance system and a “water dispenser immersion”. The confocal spinning disk is the Yokogawa mono-camera SD CSU-W1 mono-disk (50µm), the Oxxus excitation laser source has 4 wavelengths 405-488-561-640 nm. The sCMOS Hamamatsu Orca Flash 4.0 camera complements this detection equipment as well as a thermostated and CO2 OKOLAB incubation chamber with different insert sizes. An off-line station allows the analysis of images with the software ZEN blue, NIS-AR, FiJi, ICY and Imaris and to manipulate large files. A reservation system has been also set up.

Microscopy systems available @IS2M

PIC-STRA aims to support the 10 CRBS research units, as well as external teams from both the academic and private sectors. Opened in October 2020, this 350 m2 imaging platform provides users with around ten imaging systems (stereomicroscopes, wide-field and confocal single- and multiphoton microscopes with super-resolution module) for multi-scale observation, from whole small animals to sub-cellular details. It provides various solutions for the observation of fixed and living samples (videomicroscopy) and is equipped for image processing and analysis (IMARIS, Fiji/ImageJ, ICY, iLastik). The platform is part of the local RISEst network (Réseau d’Imagerie Strasbourg grand Est), is in the process of obtaining CORTECS and IBISA (STrasbourg Centre) accreditation and works closely with the other platforms in the network.

Microscopy systems available @PIC-STRA

The PI2 imaging technical platform belongs to the UPR3572 research unit: I2CT (Immunology, Immunopathology and Therapeutic Chemistry) of the IBMC (Institute of Molecular and Cellular Biology) in Strasbourg. The platform has i) sample preparation tools, ii) microscopy/imaging tools and iii) image processing and analysis tools. The platform offers workflows involving these 3 stages in order to answer the biological questions raised by scientists. The platform is part of the local RISEst network (Réseau d’Imagerie Strasbourg grand Est) and works in close collaboration with the other platforms in the network.

Microscopy systems available @PI2

The platform is attached to ITI Neurostra. It is located at the Institut des Neurosciences Cellulaires et Intégrées (INCI) and managed administratively by the CNRS UAR 3156 headed by P. Darbon.
The expertise of the in vitro imaging platform applies to the study of biological materials, tissues and isolated cells on a structural and ultrastructural scale. It extends to the immunocytochemical visualisation of molecules and the phenotypic detection of gene expression.
The platform is involved in a wide variety of interdisciplinary projects (biology, chemistry, agronomy, medicine). Around two-thirds of users come from the CNRS INCI UPR3212 (6 groups), with the remainder coming from other Strasbourg institutes (IBMC, IBMP, ISIS, IPHC, ECPM, CRBS, etc.).
A number of developments have taken place in recent years, including cryofixation (Wohlwend system) and super-resolution microscopy (Stellaris 8 confocal microscope with a STED module).

Microscopy systems available @Plateforme Imagerie In Vitro INCI

The microscopy facility was the first technology platform established in the IBMP in 1998. Its scientific programme aims at understanding the expression of plant or animal genes over space and time at various levels. Microorganisms or biomaterials studied by partner research units are other topics of interest. Our facility follows official guidelines for « Plates-Formes Technologiques du Vivant » and has received RIO 2001, 2004 and 2006 labels. It’s part of the larger PIC.sc Strasbourg Centre Cell Imaging Facility that allows sharing devices and knowledge from several research units from CNRS, INSERM, UNISTRA. Our missions include assisting research from IBMP and partner research units, developing and implement new imaging technologies, training our user base and beyond, getting involved in microscopy education and science popularization.

Microscopy systems available @Plateforme d’Imagerie Cellulaire de l’IBMP

The QuESt imaging facility combines the microscopy resources of the Institut de Génétique et Biologie Moléculaire (IGBMC) and the Laboratoire de Bioimagerie et Pathologies (PIQ). The two laboratories are located on the Illkirch bio-campus, just 10 minutes’ walk from each other. QuESt has held the IBiSA label since 2014. QuEst offers a range of instruments for multi-scale imaging, from the molecule to the whole animal. The ICI (Imaging Center IGBMC) component located at the IGBMC specialises in imaging the dynamic processes of living organisms at the molecular, cellular and whole organism levels. Researchers can analyse, in an integrated manner, their study models at different resolutions, ranging from the finest cellular structures to the complex functioning of organs in vivo. The PIQ (Plateforme d’Imagerie Quantitative – Quantitative Imaging Platform), located in the Faculty of Pharmacy, has a specific focus on quantitative molecular microscopy methods. In addition to commercial instruments, the PIQ-QuESt develops its own state-of-the-art instruments.

Microscopy systems available @QuESt

Toulouse Réseau imagerie (TRI) platform is a large multisite platform, federates all shared imaging resources in biology lab in Toulouse Area. TRI offers, through 4 fields of expertise which are photonic microscopy, electron microscopy, cytometry-cell sorting and image processing and analysis, the visualization of biological functions, from the nanometric scale to the whole organism.

The missions of the platform are threefold:
1- To provide researchers expertise and cutting edge technology in the field of :
• Mecano-biology
• Molecular and single cell characterization
• Whole organism, including intra-vital imaging
• Image analysis, signal treatment and modelisation
2- To constitute a framework for reflexion and exchanges in this very rapidly evolving field.
3- To contribute to the formation of researchers in all methods related to microscopy.
TRI platform is ISO9001-NFX50-900 certified and IBiSA facility labeled.

Microscopy systems available @TRI

Located in Rouen, PRIMACEN, the cell imaging facility of Normandy (1000 m2) is integrated since 2022 in HeRacLeS (Univ. Rouen Normandy, Inserm US51, CNRS UAR2026). To achieve its service offering and R&D objectives, PRIMACEN proposes a continuum with 3 pillars thanks to complementary skills in cell biology-histology, instrumentation-quality control and data processing. In tight connection with animal (rodent/marine models) and plant facilities, the first transversal pillar involves living and fixed sample preparation including labeling, for advanced light and electron microscopies. Then, appropriate resolutions are proposed to users and collaborators through multi-scale and multi-modal imaging including LSM and 2P, WF/confocal micro/macroscopy, TIRFM, FLIM-STED imaging, cryo-FLIM/confocal and TEM/cryoTEM microscopies. To finally help users in their scientific projects, image processing and analysis (3D modeling, ImageJ macros, AI…) are available on highly performing working stations.

Lyon Multiscale Imaging Center is a microscopy facility opened to academic research and industry. It federates 3 imaging centers in the Lyon area and provides multi-scale imaging tools including light and electron microscopy:

• the PLATIM, located at the Ecole Normale Supérieure de Lyon and at Lyon-Sud Medical University, offers a wide range of light microscopy techniques ranging from fully automated video-microscopy, confocal and two-photon microscopy. The PLATIM accounts also for a platform of atomic force microscopy and microindentation for acquiring high resolution 3D topographies and for measuring micro/nano-mechanical properties.
• the Centre Technologique des Microstructures CTµ, located on the La Doua campus, is mainly an electron microscopy facility that handles multi-disciplinary projects at the interface between biology, chemistry and physics. the CTµ provides access to a wide range of scanning and transmission electron microscopy techniques, and masters cryomethods for sample preparation and observation.
• the East Lyon imaging center (CIQLE) is located in the Rockefeller Medical University. It provides access to wide-field and confocal light microscopy and transmission electron microscopy for the bio-medical research community.

Machines can be operated independently by the users after appropriate training. Users can also benefit partial or full assistance for their projects. LyMIC provides training sessions for groups or individuals.

The team is watching technology progress and implement new techniques to provide state-of-the-art tools.

ENS Chemistry facility gathers instruments devoted to the characterization and purification of optical probes and actuators by means of various spectrometries (UV-Vis absorption, fluorescence emission) and chromatographies (capillary electrophoresis, HPLC analytical or preparative), installed on 100 m² at ENS Chimie. Access is provided to external users with technical and conceptual assistance from the 8 permanent members involved in FBI. The originality of ENS Chimie is the close collaboration between the characterization facility making available established approaches to the biological community and a research team involved in the development of state of the art chemical technologies for the optical control and reading out of living systems. The ultimate goal is to provide access and training to these emerging techniques and methods for the realization of competitive biological projects.

Choquet team

The team develops several research topics, combining neuroscience, imaging, chemistry and protein
engineering in order to unravel the dynamics and nanoscale organization of multimolecular receptor
complexes and their functional role in glutamatergic synaptic transmission. We develop 3 main
research axes to understand the interplay between AMPA type glutamate receptor nanoscale
dynamics, synaptic plasticity and memory formation in the healthy and diseased brain: 1) dynamics
and physical-chemistry of the macro-molecular complexes of the synapse, 2) nano-scale organization
and dynamics of synaptic proteins and membrane trafficking, 3) impact of the dynamic of synapse
organization on synaptic physiology.

Groc team

We specifically investigated the trafficking and function of the NMDA receptor (NMDAR), as well as dopamine receptors, since their physiological activity is central for the maturation of networks, synaptic plasticity, learning and memory, and is currently one of the most studied targets for neuropsychiatric disorders. Indeed, a deficit in NMDAR signaling in the brain is currently proposed as the core molecular dysfunction of brain circuits in psychotic disorders. The team contribution to the field has been to demonstrate that i) these receptors are highly dynamic at the surface of neurons, ii) the dynamics is regulated by intracellular, transmembrane, and extracellular interactors, iii) their dynamics is essential for activity- and hormone-dependent synaptic plasticity and associative memory, and iv) this surface dynamics is strongly dysregulated in psychotic disorders (e.g. autoantibodies). Our discoveries were possible thanks to interdisciplinary efforts, in which for instance we developed single nanoparticle tracking in live brain tissue to define receptor behavior and the extracellular space ex vivo.

Naegerl team

The team develops and applies advanced super-resolution microscopy approaches to study the
molecular and cellular mechanisms of cell-to-cell communication and its dynamics in the mammalian
brain, using the mouse as a model system. Notably, we invented super-resolution shadow imaging
(SUSHI) to visualize the extracellular space of the brain in the living state. 

Sibarita team

It is an interdisciplinary team composed of physicists, computer scientists and bioengineers having a strong interest in biology. The team is developing cutting edge quantitative imaging and analysis techniques to decipher protein organization and dynamics with high spatial and temporal resolution and high content screening context. Recent developments include analysis software for single molecule-based super-resolution microscopy (SMLM), innovative single objective light-sheet microscopy, SMLM-based high content screening platform and correlative STED-SMLM super-resolution platform. The Quantitative Imaging of the Cell has an important academic and industrial transfer technology activity.

Giannone team

Our goal is to decipher at the molecular level the spatiotemporal and mechanical mechanisms controlling the architecture and dynamics of motile structures. Exploration of these new dimensions requires an innovative and multidisciplinary approach combining cell biology, biophysics, biomechanics, super-resolution microscopy, single protein tracking and quantitative image analysis.

Studer team

This team is a joint research lab between IINS and the company Alvéole (www.alveolelab.com). By joining their research efforts in a joint research laboratory (JRL), Alvéole and IINS expect to become key players in the development of human in vitro models for fundamental research, disease modeling and drug testing. The team develops instruments and methods to prototype and image advanced in vitro cellular models.

Perrais team

The team studies the nanostructure of exocytosis and endocytosis sites at synapses, the cellular and molecular mechanisms of endocytosis. We extend the topic to diverse synapse types (glutamate, dopamine…) and how these processes are affected in early stages of neurodegenerative diseases (synucleinopathies). We develop new probes (pH sensors) to detect exo- and endocytosis events at the highest resolution possible. We also develop fluorescence-based strategies to purify synaptosomes for cellular and molecular omics analyses.

Thoumine team

The team investigates then mechanisms of synapse assembly in neuronal culture systems with a focus on the role of adhesion molecules, using a combination of experimental approaches ranging from single molecule imaging to optogenetics and electrophysiology. Recent developments include the design of small fluorescent probes for super- resolution imaging of synaptic adhesion molecules (Chamma et al., Nat Comms 2016), the release of a simulator of single molecule dynamics for fluorescence live-cell imaging (Lagardère et al., Sci Reports, in press), and the optogenetic stimulation of phosphotyrosine pathways for the control of synapse differentiation (Letellier et al., eLife 2020).

The Developmental Biology Institute of Marseille (IBDM) is an international and interdisciplinary research institute oriented towards developmental biology and pathologies. The research activity is at the crossroads of development, neurobiology, cell biology, biophysics and genetics. The general theme of IBDM is to understand how the instructions encoded in the genome are translated to build structures (cells, tissues, organs) that perform specific functions, and how these processes are regulated and integrated in the whole organism. There are links between developmental biology and diseases such as cancer, neurodegenerative and genetic diseases. One of the priorities of IBDM is to foster interdisciplinarity through the integration of new and original approaches that create conceptual and technical interfaces. At IBDM, Lenne team aims at determining how (1) mechanical and (2) physical interactions are organized at cell surfaces in vivo and (3) how these interactions are processed to produce cell and tissue responses. To tackle these questions, we focus on two aspects of tissue morphogenesis, namely cell polarization and force transmission in fields of cells. We are using Drosophila and C. Elegans as systems to address questions (1-2) and question (3), respectively. The originality of our project relies in the integration of both physics and experimental biology to study quantitatively tissue morphogenesis.

Founded in 1976, the Centre d’Immunologie de Marseille Luminy (CIML) is a research institute internationally renowned in its discipline. From worm to man, from molecule to the whole organism, from physiology to pathology, the CIML addresses, over numerous models and scales, all fields of contemporary immunology: the genesis of different cell populations, their patterns of differentiation and activation, their implication in cancer, infectious and inflammatory diseases and the mechanisms of cell death. At CIML, Marguet team aims at understanding the role of membrane lateral dynamics and organization in T lymphocyte signaling, by analyzing the molecular interaction/association events at high spatial-temporal resolutions. A special emphasize is made at examination of the molecular dynamics in the plasma membrane to initiate and to integrate extracellular stimuli. In this context, Marguet team develops analytical methods based on the combination of single molecular sensitive detection approaches such as fluorescence correlation spectroscopy (FCS) and derivatives, of single particle tracking with optical tools allowing to manipulate the biological samples such as dynamic holographic optical tweezers.

The Mosaic group of the Fresnel Institute headed by H. Rigneault has been involved for almost a decade in developing dedicated optical instruments for biological imaging. Among other, the team has developed together with CIML the “FCS diffusion law” approach in Fluorescence Correlation Spectroscopy that has been successfully applied to the cell membrane. More recently single particle tracking using multiple targets have proved to be powerful to distinguish confinement zone at the cell membrane and Holographic Optical Tweezers shows potential application into TCR/MHC control. Phase control for micro-manipulation and imaging is an active field of research at Mosaic. Since 2002, the Mosaic group has been involved in coherent Raman microscopy and nonlinear imaging and was the first in France to build and develop a CARS microscope. One of the group world recognized expertise is in polarization resolved fluorescence and nonlinear microscopy that has proved to be able to retrieve molecular order in cell and tissue imaging. The group is now also involved in the development of nonlinear imaging using endoscopes using innovative microstructured optical fibers. Another active field of research is fluorescence enhancement at the nanoscale using metallic nano-antenna that have the ability to perform dynamic analysis on time and spatial scales unreachable with far field optics.

The team investigates morphogenetic processes in morphogenesis in Metazoans. We focus on the integration of cellular, molecular and genetic processes based on long-term 3D+time imaging of developing organisms in normal and perturbed conditions, data reconstruction and quantitative analysis. Methodological developments include: assisted microscopy based
on machine learning strategies to optimize live imaging conditions by multiphoton point laser scanning microscopy and light sheet microscopy, automated image processing and quantitative data analysis, interactive visualisation for cell lineage data validation, correction and analysis.

Expertise of the Team:

• Machine learning for assisted microscopy and image processing
• Optimization for in vivo and in toto 3D+time imaging by multiphoton point laser scanning microscopy and light sheet microscopy of developing organisms (metazoans an plants)
• Data management, image processing workflow, interactive visualisation

Ecole Polytechnique is an internationally attractive institution combining research, teaching and innovation. Laboratory for Optics and Biosciences (LOB) is affiliated with the French Research institutions Inserm and CNRS, and Ecole Polytechnique. The Laboratory employs researchers with expertise in optics, molecular and cellular biology with the aim to explore new concepts and methods. Two LOB research teams are involved in optical imaging developments co-funded by France BioImaging. Access to instruments is currently provided on a collaborative basis and will be extended within the timeframe of the project.
* “Advanced Microscopies and Tissue Physiology” (E. Beaurepaire, M.-C. Schanne-Klein, W. Supatto, G. Gallot, M. Joffre et al). LOB “advanced microscopies” pole is a leading group in nonlinear microscopy of live tissues and small organisms, and develops pioneering approaches based on multimodal multiphoton imaging (multicolor 2PEF, SHG, THG, FWM, CARS), polarimetry, light-sheet illumination (SPIM), photomanipulation, wavefront control, pulse shaping.
* “Nanoemitters and Single Molecule Tracking” (A. Alexandrou, C. Bouzigues, et al). LOB “nanoemitters” team develops innovative assays based on non-blinking lanthanide-ion doped nanoprobes and microfluidics devices to study signaling processes in live cells.

The scientific project of the BioImage Analysis (BIA) unit is to develop image analysis and computer vision tools for the processing and quantification of multi-channel temporal 3D sequences in biological microscopy. The topics are centered about the development of new algorithms for multi-particle tracking, deformable models, mathematical imaging and spatial distribution analysis. The group has produced powerful tools for spot detection and counting in real-time imaging of virus and genes, movement and shape analysis in 3D+t microscopy and cell growth analysis. These methods and algorithms have now been regrouped under the open-source and free platform Icy (http://icy.bioimageanalysis.org), which provides a comprehensive framework for extended reproducible research in bioimage informatics. They have been instrumental for the successful achievement of a large number of collaborative biological projects.

Institut Curie is active in image databases and management. The PICT imaging facility is engaged since 2011 with the Strand Life Sci. company in the development of the CID (Curie Image Database)/iManage (supported by Paris-Centre Node). The CID is linked to the “Curie Image Data center” (2x 100Tb Storage equipment and cluster for image processing and analysis). Since December 2013 CID is open to all FBI users of the PICT, under demand and common rules of imaging platforms (web client). iManage is the commercial version (with licensing), offering support to labs, to install and adapt CID on their own microscopy, at their own sites. Plugins to access the CID from Icy (Institut Pasteur) are developed and published on the central repository of Icy. An interface to interoperate with the servers at Institut Curie is under development. Integration of software developed in collaboration with Inria-Serpico (http://mobyleserpico.rennes.inria.fr/) such as ND-SAFIR, Hullkground are now integrated in the CID for automated treatment. Institut Curie, specifically with Serpico’s Team@Inria-Rennes, also develops new algorithm solutions for dynamic events detection and classification, sub-diffraction light microscopy and CLEM approaches.

The Serpico team provides computational methods and mathematical models to automatically extract, organize and model information present in temporal series of images as they are obtained in multidimensional light and cryo-electron microscopy. In the field of membrane traffic, Serpico addresses the following themes in close collaboration with Curie Institute: image superresolution and image denoising to preserve cell integrity (photo-toxicity vs exposure time), information extraction from images and videos in multidimensional microscopy for molecular interaction analysis, spatiotemporal modeling of molecular species and multi-scale architectures, computational simulation and modeling of membrane transport at different scales. In collaboration with UMR 144 and PICT at Institut Curie, the members of Serpico participate in several joint projects (PhD and post-doc supervision, industrial contracts…). They have proposed user-friendly algorithms for processing 3D or 4D data. Other projects are related to instrumentation in microscopy including computational aspects (SERPICO@Mobyle web service) and data management (CID iManage) on the reconstruction and enhancement of images related to subdiffraction light microscopy and multimodal approaches.

We develop single-molecule and advanced microscopy methodologies to investigate the mechanisms underlying DNA segregation and remodeling in live cells.

Our current research projects:

Our research aims at characterizing macromolecular complexes governing major biological processes, focusing on transcription regulation, signaling and remodeling of biological membranes. To achieve these goals, we develop, combine and use advanced single molecule biophysical methods (such as atomic force and fluorescence microscopies), as well as DNA nanotechnology.

Research:

Structure and dynamics of nucleoproteic and membrane assemblies

• Prokariotic transcription termination
• Bacterial adaptation to environmental changes
• DNA nano-engineering

Structure and dynamics of membrane assemblies

• Remodeling and partition of membrane components
• Nanomechanics of cancer metastatis
• Mechanisms of nuclear pore complex assembly
• Structural dynamics of single metabotropic glutamate receptors
• Methodological developments

The SIMS Group (Signal, Image and Sound) develops its research in the field of statistical signal and image processing. Tackling ill-posed inverse problems such as deconvolution and diffraction tomography is one of our main fields of expertise. We also focus on computational imaging issues, relying on mathematical tools from both optimization and Bayesian simulation fields. The optimal co-design of computational imaging systems is among our growing topics of interest, with the viewpoint of information theory. In the area of fluorescence microscopy, we investigate the superresolution capacity of random illumination microscopy, in collaboration with Institut Fresnel (Marseille) and the Centre for Integrative Biology (Toulouse).

Expertise of the Team:

• Inverse problems
• Computational imaging
• Machine learning

The team aims at developing techniques and methodologies in fluorescence microscopy to study dynamics of protein-protein interactions and biochemical activities in live sample. Team approaches are mainly driven by methodological and technological development, its transfer and its applicability in biology to answer relevant new questions. Recently, the team is investigating the possibility to implement smart and autonomous microscopy approches to be able to develop robust and automated high content microscopy pipelines of cutting-edge methods such as F-microscopy or photoperturbative experiments.

The Imaging group within the Nanobio team is composed of 8 people, with 1 CNRS researcher, 1 MCF, 4 PhD students and 1 engineer on temporary contract. The team develop new fluorescence imaging microscopy modalities for live cell imaging, FLIM and super-resolution.

For most biological questions, it is essential to monitor in vivo and in real time the complex cellular machinery with extreme sensitivity and ultimate resolution. The fluorescence thus remains the tool of choice for specific monitoring in biology. Beyond imaging intensity to a specific location, fluorescence and its various spectroscopic properties can be used to locally probe its environment or reveal interactions. The photophysical properties of fluorophores are the basis of recent developments, which have enabled microscopy to turn into nanoscopy, exceeding the resolution limit imposed by diffraction, enabling the detection of single molecules with nanometer resolution. This level of resolution allows to consider the study of biological systems at a scale unprecedented in optical microscopy. The developments of the team aim to correlate functional and structural 3D information in different biological applications. In particular we develop different strategies to take advantage of supercritical angle fluorescence (SAF) emission and also introduced a time approach to localize single molecule in structured excitation (ModLoc)

The OV-Cytology and Imaging Team (OV-CI) is part of a multi-level IBISA plant phenotyping facility named “Observatoire du Végétal (transcriptome, proteome, metabolome, but also cell and tissue organization). It is hosted at the INRAE at the Institut Jean-Pierre Bourgin (Saint-Cyr l’Ecole) near Versailles, in Ile de France-Sud, and is associated with the University Paris-Saclay.

The OV-CI team is animated by 8 permanent people (full or part-time), with 1 INRAE researcher, and 7 INRARE engineers. The OV-CI team is mainly devoted to take up challenges in plant Imaging.
It provides new methods, protocols, or devices/equipment in BioImaging necessary to tackle the plant front science. Recent developments include a workflow for live meristem imaging, microfluidic devices for roots imaging, specific configuration for plant vertical fluorescence imaging to study tropism, sensors and growth monitoring, etc.

The ICAR Research-Team develops research themes combining the interaction and processing of visual data such as 2D, 3D, videos or nD+t image sequences and 3D meshes. The team is structured according to 3 research axes: Analysis & Processing which is focused on new low-level processing techniques of visual information, Multimedia Security which is interested in securing visual data and Modeling & Visualization which aims at representing large and complex visual data sets.

Expertise of the Team

• The Analysis & Processing axis is interested in new low-level information processing techniques to improve the information perceptible in the image and to take into account, within the same theoretical framework, the imprecise, uncertain and incomplete (the different types of error in
  visual data processing).
• The Multimedia Security axis is interested in the security of visual data. In order to ensure this security, coding algorithms are developed combining tattooing, steganography, forensics, encryption and authentication and often requiring robustness to compression.
• The objective of the Modeling & Visualization axis is to model large sets of complex data (in dimension and nature) in order to allow intuitive visualization or to manipulate these data to extract knowledge from them.

The team is concerned with developing novel optical imaging methods using scattered light. This includes methods for deep imaging in the multiple scattering regime, super-resolution and non-linear coherent and incoherent microscopies.
Expertises of the Team:

• Imaging in the multiple scattering regime
• Super-resolution
• Computational imaging for fluorescence and Raman

The team is a pioneer of Cybergenetics, which aims at controlling biological systems in real time thanks to computercontrolled feedback loops fed by real time image analysis and driven by microscopy automation. We are developing novel software solutions to enable smart microscopy applications.

Expertise of the Team

• Smart microscopy
• Dynamic control of living systems
• Microfluidics for biology

We focus on the functions of phagocytic cells in normal and infected conditions. We have developed dedicated imaging techniques to monitor with high spatial and temporal resolution the mechanims of capture and degradation by phagocytic cells, their impact on immune responses and their alterations by viral infections, which can lead to the development of bacterial co-infections or uncontrolled inflammation.

Expertise of the Team

• TIRF microscopy with dedicated analysis of phagosome closure assay in three dimensions on living cells
• TFM adapted for living and phagocytosing cells (collaboration with M Balland, LiPhy, Grenoble)
• New FRET probes to analyse receptor clustering on living cells (collaboration with J Fattaccioli, ENS and Institut Pierre Gilles de Gennes, and Jean-Maurice Mallet, ENS Paris, programme 80 PRIME CNRS and ANR 2021-2024).

The team develops high-resolution systems for nanomanipulation and imaging of single molecules, with a focus on analysis of DNA metabolic processes such as gene expression and DNA repair. Recent developments include the synthesis of molecular DNA scaffolds for the generic study of protein-protein and drug-protein interactions in real-time and at single-molecule resolution.

Expertise of the Team

• Single-molecule experimentation (single-molecule nanomanipulation, single-molecule fluorescence)
• Biochemistry
• Optics

The team develops experimental and computational imaging and modelling approaches for cell biology and microbiology, with a focus on single molecule localization microscopy, deep learning, chromatin organization and spatial transcriptomics.

Expertise of the Team

• Single molecule localization microscopy (optics and computational image reconstruction)
• Deep learning (applications to biological and medical imaging)
  RNA-FISH and quantitative analysis

Our group has strong interests in gene expression mechanisms, from transcription to translation. While we are interested in the regulation of these processes and their functional consequences, the big question that moves us is to understand how they occur in the context of a living cell.

Indeed, cells are not only the individual units where gene regulation takes place, but they are also incredible objects: if we consider RNA and proteins, a typical cell contains several hundreds of thousands of different molecular species, with some present in millions of copies per cell while others in only few. In order to function with such a high complexity within a crowded molecular environment, cells rely on two main tools: (i) chaperones specialized in the control of molecular interactions; (ii) a remarkable degree of spatial organization, which also allows a high plasticity and a high dynamics of molecules. It is to get insights into these very fundamental questions that we first developed tools to image single mRNAs in live cells. With these tools in hands, and others that we developed later, we aim at imaging the basic mechanisms of gene expression directly in living cells, thereby providing a renewed vision of these fundamental processes.

Our strategy is to invest in methodological developments to access and image new facets of gene expression, usually at the levels of single molecules. These developments are mostly focused on imaging RNA metabolism and they are guided by our current scientific questions.

Methodological developments require multidisciplinary approaches, and we have therefore developed a stable network of collaborators who complement our own expertise. This includes the groups of: (i) C. Zimmer and F. Müller (Pasteur Institute, Paris; https://research.pasteur.fr/en/team/imaging-and-modeling/), a physicist team with a great expertise in image analysis; (ii) T. Walter (Curie/Ecole des Mines; Paris; http://members.cbio.mines-paristech.fr/~twalter/), an applied mathematician expert in high-content microscopy and in complex, high-dimensional dataset analysis; (iii) O. Radulescu (Montpellier University; https://systems-biology-lphi.cnrs.fr/), a mathematician expert in modeling biological processes. More recently, we initiated collaborations with chemists to develop novel RNA probes and biosensors.

Our group works in three main areas: transcriptional and translational regulation, as well as chaperone-mediated control of molecular interactions.

The lab is focused on the algorithms and computation selected by evolution to perform biological decision-making. We address this topic with an interdisciplinary approach mixing statistical physics, Bayesian machine learning, information theory and various experimental biological setups. We are pursuing 4 research axis:

• Probabilistic pipelines and Artificial Intelligence to probe single biomolecule random walks
• Decision-making of biological system
• Amortized inference in Virtual Reality: DIVA + Genuage
• Numerical Methods for temporal networks

Expertise of the Team:

• Probabilistic pipelines for microscopy data analysis
• Virtual reality and augmented reality applied to visualisation and analysis
• Bayesian Inference, amortised inference and physics-based Bayesian induction.

The team develops deep learning models and large scale image and data analysis algorithms. Applications range from basic research in developmental biology and neuroscience to drug discovery in collaboration with the pharma industry. We provide the source code of our methods with all papers we publish.

Expertise of the Team:

• Deep learning
• Image processing & analysis
• Computational biology

The team develops tools in modeling, simulation, data analysis in cell biology and medicine. Recent developments include new super-resolution SPTs analysis.

Expertise of the Team:

• Modeling: biology at multiple scales.
• Simulation: stochastic and deterministic
• Data analysis: trajectories, EEG, electrophysiology data.

Nanomaterials nowadays occupy an indisputable place for their diagnostic and therapeutic potentialities, offering new paradigms in the field of medicine in terms of follow-up, safety and personalized treatments. Due to the complexity of the biological environment, mastering the synthesis and understanding the interplay between nanomaterials and biological entities, especially cells, is an imperative need to overcome numerous pitfalls related to toxicity, chemical instability, undesirable targeting, diagnostic artefacts and bioaccumulation, to cite only the major ones.

To this aim, we have been developing for several years multimodal nanoassemblies, originally based on small-molecule photoactive materials, self-assembled as a platform and comprising functional inorganic nanoparticles to bridge the gap from in cellulo to in vivo investigations. Their versatile fabrication offers straightforward architectural tunability, allowing us to decipher their in cellulo interactions.

IMTI team develops innovative microscopes to image unlabeled samples. Such systems achieve 3D refractive index distribution measurements at cellular level, with twice-improved resolution compared to conventional optical microscopes. We have achieved several premieres in the domain: 

2009: world-first TDM with improved resolution: 130 nm, world record at that time (Opt. Lett. 34, p. 79 (2009)). 

2010: world-first images in correlative TDM-Fluorescence microscopy (J. Biophoton. 3, p. 462 (2010)). 

2010: world-first dual transmission-reflection TDM images (J. Mod. Opt. 57, p. 740 (2010)). 

2014: world-first high lateral resolution/high longitudinal precision profilometry (Appl. Opt. 53, p. 748 (2014)). 

2017: world-first improved and isotropic resolution TDM images (≈180 nm in 3D, distinguishing absorption and refraction. This achievement was awarded the cover of Optica, April 2017 (Optica 4, p. 460 (2017)). 

2019: world-first experimental mirror-assisted TDM images (albeit in simplified configuration) (J. Opt. Soc. Am. A 36, COF1 (2019)).

The highly multidisciplinary Biophotonics team develops fluorescence-based imaging techniques and innovative probes, in order to apply them for solving key questions related to the molecular and cellular functions and interactions of viral and epigenetic proteins with their nucleic acid targets. The physicists and engineers of the team have implemented and developed home-made microscopy set-ups, which are regularly upgraded and optimized for new applications and projects. The team implemented in 2003 one of the first two-photon FLIM/FCS microscopes in France. Later, the team developed a high resolution 3D dSTORM/PALM microscope and a STED microscope with cw laser. In order to track and image UCNPs, an anti-Stokes wide-field for SPT and an anti-Stokes luminescence lifetime confocal microscope were developed. More recently, a spectral PAINT (sPAINT), a light sheet single molecule microscope and a single molecule UV microscope were developed. These instruments are accessible to the community through their integration into the platform PIQ-QuESt (IBISA).

The IPP team (Photonic Instruments and Processes, ICube laboratory) has a long-standing expertise in microscopy and coherent imaging, as well as diffuse optics for surgical guidance. In particular, the team develops full-field OCT (scanning interference microscopy), endoscopic OCT, digital holography, hyperspectral imaging and quantitative phase imaging for biomedical application. Moreover, the team has a particular interest in multimodal imaging and contrast for example spectroscopy, elastography and Doppler imaging. The team has also more recently started work on fluorescence imaging, and in particular light-sheet microscopy for high-throughput imaging of live biomedical samples, based on the single-objective Oblique Plane Microscopy technique. The strategy of the team is to tackle biomedical questions with optical imaging with a multiscale approach from cellular scale to the organ scale.

The team develops multifunctional fluorescent molecules and nanoparticles for biomolecular detection and imaging. We developed i) solvatochromic probes for ratiometric and super-resolution imaging of plasma membrane organization and cell polarity mapping, and ii) fluorogenic membrane probes operating from green to near-infrared range for imaging cells, tissues and organoids by advanced microscopy techniques. We also introduced environment-sensitive fluorogenic probes for imaging G protein coupled receptors and intracellular RNA. We also developed dye-loaded fluorescent polymeric nanoparticles (NPs) 100-fold brighter than quantum dots of similar size. They were successfully applied for single-particle tracking in cells and mice brains by intravital microscopy. Using NPs of different colors, we introduced a technique for barcoding of living cells. These NPs were also used as light-harvesting nanoantennas to prepare biosensors for cancer diagnostics and fluorescence in situ hybridization (FISH) for intracellular RNA imaging.

As part of the Integrated Structural Biology platform at the Centre for Integrative Biology (hosting the national and European Infrastructures FRISBI, Instruct-ERIC and iNEXT-Discovery), and our research group at CBI/IGBMC we have recently done a series of developments in super-resolution fluorescence microscopy to facilitate single molecule localization microscopy (SMLM) analysis with the aim of integrating structural data generated through crystallography, cryo-EM, cryo-ET and FIB towards the cellular level (Biol Cell., 2017). We developed an integrated software for image reconstruction, drift & chromatic aberration correction, co-localization, resolution estimation (SharpViSu; Bioinformatics 2016), segmentation & clustering of labeled complexes [ClusterViSu;4], including 3D analysis and segmentation of SMLM data using Voronoi diagrams [3], as summarized in a book chapter [1]. Our latest development comprises a spectral demixing method which facilitates co-localization of proteins in SMLM (https://doi.org/10.1101/2021.12.23.473862).

The team is dedicated to the synthetic development of new tools for analytical and imaging applications. We notably develop luminescent labels for microscopy imaging and bio-analytical applications such as luminescence based assays. The team has a strong background in synthetic organic chemistry applied to the design of lanthanide based luminescent labels. Thanks to the spectral and temporal signature of lanthanide elements, the probes have exceptional characteristics such as very long lived excited states, line-like emission spectra and large pseudo Stokes’ shift. The main scientific axes related to the project are the design of luminescent probes displaying upconversion at the molecular and supramolecular level and the optimization of lanthanide based nanoparticles for time-resolved luminescence microscopy. Regarding upconversion, our molecular approach is expected to remove the troubles associated with the use of nanoparticles (biotoxicity, reproducibility,…) while profiting from the spectroscopic advantages of the anti-Stokes process. For lanthanide based nanoparticles, the displacement of the excitation wavelength towards the visible region will provide strong perspectives for luminescence microscopy and flow cytometry. Our team is fully equipped for organic and inorganic synthesis as well as for the spectroscopic characterization of the luminescent objects with UV-Vis-NIR absorption spectrometers and UV-Vis-NIR emission spectroscopy (up to 1600 nm) in the steady-state and time-resolved modes.

3D printing and 3D Bioprinting applied to biodetection.
Development of 3D microenvironment as models for cell culture and cancer study.

Topics: Microenvironement for cell culture, tissue engineering, cancer diagnosis, microphysiological systems.
Skills: Microfluidics, 3D printing, Bioprinting, Self assembly, Biopatterning

Development of microfluidic confining culture chambers, in order to study the impact of spatial confinement. Adapted to mechano-biology and allowing well-defined microenvironment.

The team develops original systems to fluorescently label and track DNA (ANCHOR technology – NeoVirtech SAS) in real time at nanoscale resolution to understand physical principles underlying regulation of gene expression and DNA repair, in cellular plasticity and tumorigenesis.

Expertise of the team: Combination of cuttin-edge techniques in optical and electron imaging, material science, cell mechanics and intra-vital imaging to elucidate how phagocytes, in particular macrophages and osteoclasts, interact with the extracellular matrix, to decipher the mechanisms of macrophage 3D migration.

3D-electron microscopy approaches and correlative microscopies at different scales, from single molecules to tissues. Strong expertise in molecular imaging by cryo-EM and single particle analysis

Expertise in 3D imaging of live and fixed organoids. Drug screening tests using morphological characterization of organoid cultures as readouts. Organization of an open platform for Organoid development.

Dévelopment of the RIM technology (Random Illumination Microscopy). Development of low cost integrated optical technology.

The MAMBO team (MAthématiques pour l’iMagerie BiOlogique) is specialized in mathematics (optimization, learning, approximation, inverse problems) and in imaging (reconstruction and analysis). It has 2 main objectives: – Participate in the improvement of microscopy techniques (e.g. RIM, TIRF, SMLM, …) by refining their mathematical modeling and by building efficient reconstruction algorithms. – Participate in the analysis of the resulting images by developing (or using) tools to improve the quality of segmentation or classification in large volumes of data. In particular, it develops new artificial intelligence models to achieve its goals.

Expertise of the team: Combination of cuttin-edge techniques in optical and electron imaging, material science, cell mechanics and intra-vital imaging to elucidate how phagocytes, in particular macrophages and osteoclasts, interact with the extracellular matrix, to decipher the mechanisms of macrophage 3D migration.

Bioorganic Chemistry team (15 people) develops bioconjugation, surface modification, multi-step synthesis, to create biocompatible chemical tools to explore the complexity of living mechanisms. The team’s expertise ranges from organic synthesis to photophysics; aiming at the development and tailoring of 1) new organic fluorophores (either push-pull or ESIPT based with high Stokes shifts and solvatochromism), with a recognized know-how in cyanins, coumarins, rhodamins, indazoles or BODIPY fluorescent scaffolds, notably their NIR derivatives, for application in conventional, two photon and STED microscopy, 2) biodetection (proteins, DNA) techniques based on profluorescent or chemiluminescent probes, and 3) new chemobiology tools such as bioconjugation reactions (and click-chemistry type tools), chemical or photoreactive cross-linkers, photolabile groups. The team collaborates with industrial companies through Carnot I2C and with international collaborators through XL-Chem program.

The translational Cardiovascular Research team develops fluorescence-based 3D imaging techniques and intravital microscopy approaches, in order to apply them for solving key questions related to the molecular and cellular functions, notably related to inflammatory responses in hearts and vessels. The researchers and technicians of the team, in collaboration with PRIMACEN, have implemented and developed set-ups for macroconfocal lymphangiography, vascular thrombosis evaluations as well as light sheet imaging, which are regularly optimized for new applications and projects. The team implemented in 2016 the first cardiac lymphangiography in rodents. Later, the team developed approaches for light sheet 3D imaging. These protocols are accessible to the community through their integration into the platform PRIMACEN (IBISA).

Team 4 of the Inserm research unit 1245 aims to characterize the contribution of neurovascular dysfunction in the pathophysiology of neonatal brain lesions while keeping in mind brain immaturity. Thereby, research projects paid attention to molecular, cellular and integrated processes leading to angiogenesis defects and neurodevelopmental consequences such as vessel-associated migration. The team’s research activity is deeply committed into translational research validated by patents and clinical protocols with the objective to develop diagnosis tools and neuroprotection strategies. To reach these objectives, the research team developed mouse perinatal models of white matter injury, in utero gain and loss of functions, and environmental models of fetal toxicity such as FASD. Endothelial and neuro-vascular dysfunctions are apprehended in the developing brains and retinas by imaging approaches (light sheet, FLIM-STED, time-lapse migration, in situ zymography, OCT) on ex vivo tissues (brain organotypic slices, cultured retinas). Image processing and analysis through notably machine learning is developed in partnership with several members of the Normandy FBI node.

The GlycoMEV team (research axis 3) aims in understanding the biosynthesis and secretion of glycoproteins with a special focus on N-glycosylation in microalgae models. As in all Eukaryotes, N-glycosylation occurs in the endoplasmic reticulum and the Golgi apparatus, where numerous enzymes act step-by-step to build and transfer oligosaccharides named N-glycans on secreted proteins. To decipher the sequence and regulation of these events, it has recently developed cryopreparation methods and imaging technologies (i.e. TEM) enabling the characterization the microalgae cell morphology and to analyse the subcellular localisation of distinct molecular actors. Furthermore, the team has acquired expertise in molecular biology techniques specific to these models allowing the generation of strains expressing various proteins fused to fluorescent reporters. They have also developed strategies based on confocal/FLIM imaging to characterize the targeting mechanisms responsible for the localization of glycoenzymes in the Golgi apparatus

The “tPA and neurovascular disorders” team within the UMR-S U1237 – INSERM UNICAEN and GIS Blood and Brain @ Caen-Normandie Institute” (BB@C – INSERM – UNICAEN- CHU CAEN) is focused on a better understanding of the physiopathology of thrombosis/ischemic neurovascular disorders by developing and applying advanced methods (from molecular to multimodal cell and in vivo imaging). Mainly dedicated to live-cell imaging, we will provide access to dynamic imaging, super-resolution microscopy (STED). We developed innovative neuroimaging tools such as biocompatible nanoparticles, multimodal probes to detect vesicles trafficking, autophagy, neuronal activity, inflammation in the field of vascular disorders. Additionally, we used multiphoton imaging to observe in vivo vascular events including inflammation, diapedesis of immune cells within the parenchyma as well as microthrombosis. Functional ultrasound localization microscopy enables us to assess brain-wide neurovascular activity at a microscopic scale.

SEBIO is a research unit specializing in aquatic ecotoxicology with a significant experience in the development and validation of effect-based tools for monitoring and predicting environmental impact of chemicals. As experts in marine biology, SEBIO develops in vivo whole aquatic organism imaging approaches aimed at achieving breakthrough in methods and physiological data in seawater shrimps, bivalves and fish larvae. By using light-sheet microscopy, multi-modal imaging and 3D-reconstruction, these projects will decipher fine structures and detailed homeostatic regulations, in particular related to vascular anatomy, immune cell migration and blood/hemolymph tissue perfusion, regarding infection disease, intertidal cycle, osmotic stress and blood withdrawal for laboratory sampling. In relation with INERIS, technological transfers to European environmental agencies and water monitoring programs will rely on a network of reference laboratories, research centers and organizations for environmental risk assessment.

The translational Cardiovascular Research team develops fluorescence-based 3D imaging techniques and intravital microscopy approaches, in order to apply them for solving key questions related to the molecular and cellular functions, notably related to inflammatory responses in hearts and vessels. The researchers and technicians of the team, in collaboration with PRIMACEN, have implemented and developed set-ups for macroconfocal lymphangiography, vascular thrombosis evaluations as well as light sheet imaging, which are regularly optimized for new applications and projects. The team implemented in 2016 the first cardiac lymphangiography in rodents. Later, the team developed approaches for light sheet 3D imaging. These protocols are accessible to the community through their integration into the platform PRIMACEN (IBISA).

We possess unique Brillouin microscopes  for imaging different cells and tissues at various scales:

• VIPA-based interferometer functioning at 660nm for low phototoxicity. Enclosed in an environmental chamber for temperature and CO2 control. Applications include single cells, spheroids and more complex tissues/organisms. Possibility of 3D imaging.
• Standard interferometer for application to mineralized tissues (teeth, bones, shells) and plant epidermises with sub-mm resolution, possibility to reconstruct dispersion curves for analysis of mechanical anisotropy.

We also possess the expertise to analyze, process and model the Brillouin data to obtain information on the mechanical properties (including anisotropy). Home-made microfluidic devices to mount the samples in special conditions (perfusion, mechanical compression, osmotic shocks…) can be engineered with the input of the Biophysics team. Standard microscopy is also available at the ILM (spinning disk, time lapse…)

Publications

• Giulia Guerriero, Alexis Viel, Veronica Feltri, Alice Balboni, Guqi Yan, Sylvain Monnier, Giovanna Lollo and Thomas Dehoux, “Predicting nanocarriers’ efficacy in 3D models with Brillouin microscopy”, Nanoscale 15, 19255 (2023)
• Guqi Yan, Sylvain Monnier, Malèke Mouelhi, and Thomas Dehoux, “Probing molecular crowding in compressed tissues with Brillouin light scattering”, PNAS 119, e2113614119 (2022). 
• Laura Bacete, Julia Schulz, Timo Engelsdorf, Zdenka Bartosova, Lauri Vaahtera, Guqi Yan, Joachim Matthias Gerhold, Tereza Tichá, Camilla Øvstebø, Nora Gigli-Bisceglia, Svanhild Johannessen-Starheim, Jérémie Margueritat, Hannes Kollist, Thomas Dehoux, Scott A. M. McAdam, and Thorsten Hamann, “THESEUS1 modulates cell wall stiffness and abscisic acid production in Arabidopsis thaliana”, PNAS 119, e2119258119 (2022). 
• T. Lainović, J. Margueritat, Q. Martinet, X. Dagany, L. Blažić, D. Pantelić, M. D. Rabasović, A. J. Krmpot, T. Dehoux, “Micromechanical imaging of dentin with Brillouin microscopy”, Acta Biomater. 105, 214-222 (2020)
• J. Margueritat, A. Virgone-Carlotta, S. Monnier, H. Delanoë-Ayari, H. C. Mertani, A. Berthelot, Q. Martinet, X. Dagany, C. Rivière, J.-P. Rieu, and T. Dehoux, “High-frequency mechanical properties of tumors measured by Brillouin light scattering”, Phys Rev. Lett. 122, 018101 (2019)

The Liphy R&D team has a long experience in collaboration, joint-development and consulting in optic projects between physics and biology labs. Recently, our team developed a robust method to obtain absolute values of FRET from any epifluorescence microscope. The QuanTI-FRET method calibrates the experimental system and the fluorophore pair, allowing for absolute FRET efficiency and stoichiometry measurements in living cells. Current work focuses on skipping the calibration step with specific samples, hence offering a direct calibration on the sample of interest. This project has been partly funded by the SATT Linksium, an intellectual property has been registered on the software part and discussions are ongoing with private partners.

Another part of the team’s expertise is reflection interference contrast microscopy which allows the measurement of distances with nanometric precision in the vicinity of a reference surface, and to assess the surface functionalization in situ without staining (quantification, quality control). The combination with force application techniques such as flow chambers permits probing biomechanics through the simultaneous control of the force (applied) and the distance (measured). This is essential to study adhesions forces of different organisms: from bacteria to immune cells. The dedicated microscope is fully automated with temperature control, and the PI is currently working on the development of a user-friendly interface for biology-oriented projects. 

Both developments are unique in France.

Since 2010, we have developped and implemented numerous methods in optogenetics, and even chemogenetics, in order to have access to dynamics and reversible perturbations of key biological functions such as cell adhesion, cell signaling, transcription factors, inflammation, functions of immune cells and even metabolism. Our lab proposes to share this expertise to the FBI users through access for consulting and even direct collaboration. In the Rhonalpin node, our lab is focused on the coupling between optogenetics, biosensors and metabolism imaging through FLIM imaging. Indeed, we have developed a TIRF microscope presenting a module of FastFLIM imaging. Through the use of dark acceptors, this technology allows the users to extend the possibilities in terms of combining optogenetics, biosensors, metabolic imaging through ratiometric probes and classical multicolors TIRF imaging. Moreover, TIRF imaging allows long term live imaging with low levels of photoxicity. This is essential for metabolic imaging and preserving photon budget for FLIM imaging. In the future, this system will be coupled with a module of evanescent field patterning (EFP) in order to have a specific TIRF-mode illumination of only  a region of interest (microm scale).

Publications

1- An optogenetic approach to control and monitor inflammasome activation. Julien Nadjar, Sylvain Monnier, Estelle Bastien, Anne-Laure Huber, Christiane Oddou, Léa Bardoulet, Gabriel Ichim, Christophe Vanbelle, Bénédicte Py, Olivier Destaing*, Virginie Petrilli*. Recently accepted in Science Signaling. bioRxiv 2023.07.25.550490; doi: https://doi.org/10.1101/2023.07.25.550490

2-Optogenetic control of YAP cellular localisation and function. Toh PJY, Lai JKH, Hermann A, Destaing O, Sheetz MP, Sudol M, Saunders TE. EMBO Rep. 2022 Sep 5;23(9):e54401. 

3-Control of SRC molecular dynamics encodes distinct cytoskeletal responses by specifying its signaling pathway usage. Kerjouan A, Boyault C, Oddou C, Hiriart-Bryant E, Pezet M, Balland M, Faurobert E, Bonnet I, Coute Y, Fourcade B, Albiges-Rizo C, Destaing O. J Cell Sci. 2021 Jan 25;134(2):jcs254599.

4-β1A integrin is a master regulator of invadosome organization and function. Destaing O, Planus E, Bouvard D, Oddou C, Badowski C, Bossy V, Raducanu A, Fourcade B, Albiges-Rizo C, Block MR. Mol Biol Cell. 2010 Dec;21(23):4108-19.

Over the last ten years, the joint effort of the platform PLATIM and the laboratory of Plant Development and Reproduction (RDP) has developed a strong and recognized expertise in the imaging and quantification of cell mechanical properties in living plant issues. Specific pipelines have been developed to map and quantify:

1. Wall mechanical properties through direct measurements of key parameters such as wall stiffness, adhesion, and resistance to piercing, using AFMs and nano-indenters 

2. Cell hydrostatic pressure through stiffness measurements made using AFMs or nano-indenters and the subsequent application of physical models but also through the development of a nano-pressure probe (in progress).

In addition of the originality of these technological approaches in mecanotransduction, the interest of our R&D teams is to develop and support their evolution directly into the environment of an IBiSA core facility. Thus, the R&D Team PLATIM/RDP is one of the rare facilities in France to provide different and integrated mechanical evaluations for both plant and animal multicellular organisms.

Publications:

• Bauer, A., Ali, O., Bied, C., Boeuf, S., Bovio, S., Delattre, A., Ingram, G., Golz, J.F. & Landrein, B. Spatiotemporally distinct responses to mechanical forces shape the developing seed of Arabidopsis. (2024) EMBO J. https://doi.org/10.1038/s44318-024-00138-w
• Creff, A., Ali, O., Bied, C., Bayle, V., Ingram, G. & Landrein, B. Evidence that endosperm turgor pressure both promotes and restricts seed growth and size. (2023) Nature Comm. https://www.nature.com/articles/s41467-022-35542-5

The development occurring in the platform MEC is tighly associated with joint effort with the “platform of nanocaractérisation” (PFNC) from CEA, leading to have a unique expertise in 3D electron microscopy of large tissue volumes reaching tens of thousands of cubic micrometers at the nanometer scale. This is essential to analyze rare events and true 3D morphological details in neurobiology to study finely cell connectivity, the characterization of membrane structures in chloroplasts from algae and plants, or to study nanoparticles toxicity. Indeed, EM stacks are acquired with much less efforts than by serial sectioning and in a relatively short time (preparation of the sample 3 days, acquisition of the stack 1 day due to high quality stabilization of the samples) new biological questions of interest to the FBI community can be addressed. 

Moreover, we have developed a unique pipe-line of analysis of large stacks of 3D EM images. This pipe-line is composed of segmentation steps followed by Ilastik approach for recognition of objects of interests coupled with a serie of home-made plugin specific for quantification. As requested by the network of French electronic microscopists, we are actively collaborating to make available our process to make this pipe-line available for others colleagues and especially the FBI users, as requested by the network of French electronic microscopists. Our work allowed us to develop international collaboration with Andrea Volterra, a world-class leader in the field of FIB-SEM in Neurobiology.

Transcriptomic heterogeneity of cells in organisms over time and space necessitates state-of-the-art technologies to access this variability and its dynamics in situ. Hence, the Spatial-Cell-ID facility aims to enhance the spatial resolution of MERFISH for pinpointing transcripts of specific genes (ranging from a few to thousands) in situ, achieving cellular and subcellular precision over time. To achieve this, we employ a microfluidics device enabling multiple rounds of hybridization (smFISH) synchronized with an advanced 3D STED microscope, providing precise spatial localization of RNA spots with a resolution of 50 nm. This technology is tailored for whole-mount samples across diverse organisms, spanning from mammalian cells to invertebrate systems such as Drosophila and C. elegans. Supported by EquipEx+ funding, Spatial-Cell-ID will be nationally accessible through the LyMIC core facility. Our facility complements commercial systems focused on imaging larger samples at lower 2D resolution. Our bespoke solution not only surpasses current state-of-the-art capabilities but also maintains its position at the forefront of technology, with potential future integration of genomic and spatial proteomic techniques.

Publications

Bouchet M., Urdy S., Guan W., Kabir C., Garvis S., Enriquez J. A simple smiFISH pipeline to quantify mRNA at the single-cell level in 3D. (2023). STAR Protocols. Volume 4, Issue 2, 2023, 102316.