Development of microfluidic confining culture chambers, in order to study the impact of spatial confinement. Adapted to mechano-biology and allowing well-defined microenvironment.
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
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.
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 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
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 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
NeurImag is a service and R&D Cell and Tissue imaging core facility of the Institute of Psychiatry and Neuroscience of Paris (IPNP, INSERM U1266 – Université de Paris) since October 2017.
It is divided in :
- Imaging services using photonic microscopy,
- Image analysis service providing « home made analysis solutions » through programming using either Icy, iLastik, ImageJ and/or MatLab.
- Primary cell culture services that provide neuronal cells, organoïds and tissue to solve biological questions through live and super-resolution imaging.
To groundwork our scientific operation, the daily management is divided into three interconnected services (Sample preparation, Imagingand Analysis) under the single management of a scientific director, Lydia Danglot (Inserm, “Chargée de Recherche Hors Classe”, Ph.D. in Neurosciences). Engineers performing technological operations are Sylvain Jeannin for Advanced Imaging, Philippe Bun for Data Analysis service, Laurianne Beynac for in vivo functional imaging and David Boulet for the Sample preparation and Primary Cell Culture service.
NeurImag manages 9 light microscopy systems for (A) the observation of biological macromolecular structures (full field) with very high spatial (3 confocal, 1 Airyscan confocal, 1 super-resolution SIM and 1 3D-STED and STED-FCS) and temporal (spinning-disk, 2 photons imaging, Light-sheet imaging) resolution or high molecular localization precision (2 super-resolution microscope for PALM, SPT and 3DSTORM) correlated respectively with high speed Airyscan or swept-field confocal (SFC), (B) monitoring macromolecular dynamics and interaction within cells and/or living organism with minimal phototoxicity (full field, spinning-disk and fast Airyscan confocal) and (C) spectroscopy approaches for monitoring intracellular and molecular dynamics using light manipulation (spinning-disk with laser ablation and FRAP addons) and fluorescent lifetime decay measurements (fast-FLIM confocal) respectively. NeurImag also provides 3 workstations directly connected to microscope computers for image processing and data analysis using free- and commercial software such as ICY, ImageJ, Huygens, iLastik, Imaris, Neurolucida, Volocity, Zen Blue (Zeiss), SRX (Bruker) and MatLab.
The NeurImag cell and tissue culture services offers a unique service for biological sample preparation ranging from cells (primary culture of hippocampal/cortical neurons or glial cells, organoïdes, cells lines and tissues) with the optimization of immunofluorescence labeling or transfecting protocols to the management of human biological samples from the Pathology department of GHU-Paris/Sainte-Anne Medical Center, located on the same campus. The service is equipped with 6 biosafety level2-culture rooms including 15 Class II A2 biosafety cabinet, 18 CO2 incubators, 6 bench top centrifuge, 5 light microscope and one fluorescent microscope for a quick cell culture/transfection check. As an example, internal and external teams can buy weekly, plates of neurons seeded on glass coverslips to directly proceed for imaging or transfection. Light-sheet microscope is in L2 environment and can welcome human and IPS samples.
Since Neuroscience is a major research topic at IPNP, the NeurImag Imaging Facility addresses imaging techniques related to issues faced in this field. Imaging techniques currently available cover a wide range of spatiotemporal resolution, ranging from video-microscopy, super-resolution imaging (PALM, STORM, STED 3D and SIM), sample preparation and quantitative image analysis. If you wish to discuss about the potential and decisive contribution of imaging techniques to your research project, do not hesitate to contact us.
List of system and services available
- Deconvolution widefield microscopy
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Videomicroscopy and slide scanner system
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Laser scanning confocal microscopy (LSCM) + Gated detectors + White Light Laser
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Spinning disc confocal system +Fast 2 colors GEMINI moduleSwept-field confocal system
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Total internal reflection fluorescence microscopy (TIRF)
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Fluorescence Correlation Spectroscopy (FCS)
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Fluorescence Cross-Correlation Spectroscopy (FCCS)
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Fluorescence lifetime Imaging microscopy (FLIM)
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Fluorescence resonance energy transfer (FRET)
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Fluorescence recovery after photobleaching (FRAP)
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2 photon in vivo imaging
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Light sheet microscopy for live (organoids, Zebrafish) or fixed samples
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Laser ablation and Photon uncaging
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Structured Illumination Microscopy (SIM)
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Photoactivated localization microscopy (PALM)
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STimulated Emission Depletion microscopy (STED) +/- Fluorescence Correlation Spectroscopy module
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Stochastical Optical Reconstruction Microscopy (STORM) with 3D biplane module
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Primary cell culture preparation + optimization on request/demand
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Sample preparation (transfection, Immunostaining, 3D cultures)
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Image Data Processing and Analysis +custom made solution
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Workstations for data analysis (commercial softwares)
The CeDRE team, led by Jacques Pécréaux (CRCN CNRS) is multidisciplinary, teaming up biologists, physicists and mathematicians. They are interested in mechanical aspects of cell division to understand its dynamical aspects and ultimately the extra-ordinary faithfulness of mitosis. Therefore, the team finely quantifies biological phenomena to recapitulate the results using physical models. In particular, the team performs advanced data analysis and modelling. It includes the fine tracking of spots like the tips of the microtubule, or super-resolution positioning of the centrosomes at a high frame rate (tens of frames per sec.). The obtained tracks are analysed statistically (for example, to classify the dynamics, reduce the dimension using PCA) or using Fourier spectral analysis to fingerprint the underlying mechanisms. To take full advantage of this data analysis, we model the sub-cellular mechanics during mitosis, in particular, the one grounded on microtubules, molecular motors and associated regulators on the one hand. On the other hand, we use agent-based simulations, especially cytosim, to screen for the variety of behaviours without solving the analytical equations. Finally, the team contributes to developing autonomous microscope by enslaving the driving of the microscope to on-the-fly analysis of acquired images, in particular using machine and deep learning.
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
We develop single-molecule and advanced microscopy methodologies to investigate the mechanisms underlying DNA segregation and remodeling in live cells.
Our current research projects:
- DNA organization and segregation in bacteria
- Eukaryotic DNA structure
- Molecular Motors
- Single-molecule & advanced optical microscopies