Data Management, Processing and Analysis

Data Management, Processing and Analysis

In the life cycle of a biological data, which goes from its acquisition to the formatting of results, the processing and analysis constitute a key step to making sens of the data. Analysis uses a set of mathematical and statistical operations whose goal is to improve the quality of the data and to extract relevant information.

Whether the data is acquired on or off site, the data management and analysis platform offers a set of skills and makes available to users computing and software resources covering the fields of processing, analysis, instrumental development and mathematics/statistics. It is therefore possible to propose targeted offers ranging from the reconstruction of raw data to their exploitation. Depending on the user’s request, dedicated staff will evaluate the needs and propose an adapted service ranging from support to autonomy.

EXPERTISE

EQUIPEMENTS

The platform relies on a set of analysis stations with sufficient computing power and high-performance graphics cards for the analysis of large volumes of data, the modeling and the visualization of very high-resolution 3D & 4D images using the most powerful software available at the moment.

3  Imaris (Bitplane) licences

  • 3D & 4D modeling, 2D & 3D tracking

2 Amira (ThermoFisher) licences 

  • 3D modeling, 3D & 4D image processing

1 Vision 4D (Arivis) licence

  • 3D & 4D modeling, pixel & object classification
iLastik Machine Learning

CONTACT

Mathieu Vigneau

Research Engineer

The platform's engineer evaluates the user's request and proposes the most suitable solution to his needs:

  • Advice and support of the user at the various stages of data processing and analysis
  • Hardware and software use support
  • Software training until users become autonomous
  • Development on request of specific scripts for the processing and analysis of large volumes of data
  • Support in the implementation of research and development projects in the form of collaboration

Cytometry

Cytometry

The platform allows the phenotyping of cells through the detection and identification of membrane and intracellular molecules. In order to purify a cell subpopulation, we also perform high throughput cell sorting, based on multi-parametric cell analysis. These subpopulations can then be studied for different parameters (transcriptome, proteome, in vivo transfer...).

RESTORE cytometry platform is a partner of the Toulouse Imaging Network Platform (TRI). It is certified ISO 9001 version 2015. The platform has a cell sorter, two analyzers and an image cytometer and offers a strong expertise on cells coming from solid tissues (adipose tissue, muscles, gums…). All the equipment is integrated in a L2 containment area.

The use of the equipment is open to research teams from public and private institutions, subject to acceptance of the projects by the manager.

expertise

Flow cytometry is a technique for the individual, quantitative and qualitative characterization of particles suspended in a flow.

Fortessa X20 (BD biosciences)

The Fortessa X20 is equipped with 5 lasers: 488nm blue, 405nm violet, 355nm UV, 640nm red, 561nm yellow-green, capable of reading 19 parameters. The Fortessa on the platform is also equipped with a 96-well plate changer. The platform is equipped with DIVA, Flowlogic and Kaluza analysis softwares, which can be used on site under reservation conditions.

MACSQuant 10 (Miltenyi)

The MACSQuant is equipped with 3 lasers: 405nm, 488nm and 640nm, which can read 10 parameters. This cytometer is also equipped with a plate changer, a marking and automatic dilution module. It allows to standardize experiments.

Cell sorting by flow cytometry allows the isolation and recovery of one or more cell subpopulations according to size, granulority and fluorescence criteria. For example, we are able to sort cells representing 0.1% of the vascular stromal fraction of adipose tissue.

ARIA FUSION
ARIA FUSION

ARIA FUSION

The sorter present on the plateforme is an ARIA Fusion (BD Biosciences) under sterile environment equipped with 4 lasers: 561nm yellow-green, 407nm violet, 633nm red, 488nm blue, able to read 18 parameters.

The sorter present on the tray is equipped with the following accessories:

  • 4 way sorting
  • sorting in tubes of 1.5ml, 2ml, 5ml, 15ml
  • 70µm, 85µm, 100µm nozzle
  • refrigeration module at the sorting’s inlet and outlet.

Sorting is carried out by the platform staff as a service at the request of the research teams and after acceptance by the platform manager.

Flow cytometry imaging is a technology that allows the visual power of microscopy and the statistical rigor of flow cytometry to be combined in a single instrument.

Image Stream X
Image Stream X

Image Stream X

The specific applications of the Image Stream X are multiple, we can mention in particular :

  • morphology analysis,
  • internalization,
  • cell signaling and molecular translocation,
  • co-localization, cell interactions, cell cycle, apoptosis,
  • spot counting,
  • autophagy and cell death.

The platform has an imaging flow cytometer: Image Stream X Mark II (Merck-Luminex). This apparatus is equipped with 3 objectives (20x, 40x and 60x), 3 lasers (405nm, 488nm and 642nm) and a camera and can therefore read 6 parameters simultaneously.

The image files consisting of tens of thousands of cell images can be routinely analyzed using the IDEAS software: analysis of up to 500 morphometric and photometric parameters for each cell image using patented algorithms. The software is present on the platform and can be used under reservation conditions.

Acquisitions are carried out by the platform staff as a “service” at the request of the research teams and after acceptance by the platform manager. Analyses can be carried out with the help or under the supervision of the platform personnel, in the form of a service or collaboration.

CONTACT

Marie-Laure Renoud 2

Marie-Laure Renoud

Engineer

The platform's engineer evaluates the user's request and proposes the most suitable solution for his needs:

  • providing advice in the development of multi-color panels,
  • providing training to users,

  • assisting users in the analysis and interpretation of their results,

  • assisting users in the implementation of research and development projects, in the form of collaboration.

Pour les utilisateurs réguliers : Réservation directe

Imaging

Imaging

One of the challenges in biological imaging is to visualize biological structures and living organisms in three or four dimensions. CERT has a range of instruments and expertise to perform this type of analysis.

expertise

One of the challenges in biological imaging is to visualize biological structures and living organisms in three dimensions and in depth. CERT has the instruments and expertise to perform this type of analysis.

TA-segmentation2: 3-dimensional reconstructions of the whole inguinal adipose tissue of mice.

SPIM

A new microscope based on a light sheet excitation, the SPIM (Selective Plane Illumination Microscope),  allows rapid optical sectioning in 3D and in depth on living samples and offers multi-views acquisition opening new perspectives in biological observation. This unique equipment is now available to the entire scientific community

MacroSPIM

The MacroSPIM, a macroscope based on a light sheet excitation, makes it possible after optical clearing to visualize tissues and whole organs by rapid in depth 3D optical sectioning. This novel approach offers multi-view acquisition possibilities and opens up new perspectives in biological observation. This unique equipment, based on developments carried out in Barcelona (IRB), is now made available to the entire scientific community by CERT.

Confocal biphotonique

Finally, CERT is also equipped with a biphotonic confocal microscope.   The use of an infrared laser for the excitation using higher wavelengths than in conventional fluorescence microscopy allows to penetrate more deeply into living samples and thus allows 3D and deep acquisitions.

Example videos made with these instruments

Spectral imaging coupled with linear unmixing is the technique of choice for detecting and differentiating fluorophores in a biological sample, even in situations where spectral overlap exists. Autofluorescence can also be treated as a separate fluorophore to resolve from the signal of interest.

Confocal microscopy

CERT has a confocal bi-photonic AxioExaminer microscope (LSM 880 Zeiss) equipped with a pulsed Ti/Sa (Chohérent) laser (NLO) and 3 visible lasers. 

Using the QUASAR technology, this microscope offers the great advantage of simultaneous spectral detection in a single scan. It is possible, in a single acquisition, without loss of time and without space-time shift, to acquire and perfectly separate more than 4 fluorochromes whose spectra overlap.

  • This technology also makes it possible to eliminate autofluorescence signal generated by tissues.
  • The NLO system also gives access to the second harmonic generation (SHG) signal, thus allowing to reveal the organization of collagen fibers of the extracellular matrix or the striations of skeletal muscles without labelling.
  • This equipment also allows to perform 3D and 4D imaging in vivo in small animals.

This technology makes it possible to follow living organisms over time with cellular resolution.

Videomicroscopes, Biphotons, SPIM

Real-time microscopy consists of imaging living samples over time with cellular resolution while maintaining temperature and respiratory conditions compatible with the survival of the samples. Samples can be cell cultures or whole organisms (mice, fish, drosophila), including tissue explants and spheroids.

We propose a variety of equipment : videomicroscopes, Biphotons, SPIM, that are all capable of Real-Time imaging. These equipments are coupled with specific modules that control the survival parameters of the samples (thermostatic chambers, CO2, anesthesia, respiratory and small animal monitoring systems).

CONTACT

Corinne Barreau

Engineer
laetitia.pieruccioni

Laetitia Pieruccioni

Engineer

The platform engineers evaluate the user's request and propose the most suitable solution for his needs by :

  • providing advice in the development of multi-color panels,
  • ensuring the training of users,
  • assisting users in the analysis and interpretation of their results,
  • assisting users in the implementation of research and development projects, in the form of collaboration.

Mechanobiology-AFM-BioFab

Mechanobiology
AFM BioFab

The MecanoBioFab develops original technological tools to support the institute's researchers in both the field of mechanical properties measurements on cells and tissues and in the field of manufacturing 2D and 3D supports for the elaboration of in-vitro models.

The AFM Mechanobiology platform develops new automated approaches to acquire the mechanical properties of cells, cell populations and tissues.

expertise

Previously confined to the field of physics, the AFM is gradually becoming more and more specific to biological applications. This instrument allows to measure with nanometric precision not only the topography of a living or inert surface but also their mechanical properties (rigidity, elasticity, adhesion, etc.).

NanoWizard® II from JPK (Life Science Version) coupled to a Zeiss inverted fluorescence microscope .

Bruker Catalyst Bioscope AFM coupled to a Nikon inverted fluorescence microscope

NanoWizard® II de JPK (Life Science Version) couplé à une microscope inversé à fluorescence Zeiss

With 3D manufacturing and 2D surface printing tools, the BioFab offers all the tools to integrate in-vitro cultures of organoids or artificial tissues with innovative physical supports or sensors. The proposed expertise aims to facilitate access to technological innovations from FabLab and technology centers.

CONTACT

Childerick Severac

PhD - IR CNRS

The platform engineer evaluates the user's request and proposes the most suitable solution for his needs. He :

  • provides advice in the design of experiments,
  • trains the users,
  • assists users in the analysis and interpretation of their results,
  • assists users in setting up research projects through collaboration with CERT.

Genomics & Transcriptomics

Designed by kotkoa (DNA) and starline (background) / Freepik. Modifié

Genomics &
Transcriptomics

Our platform is specialized in genome and transcriptome studies and offers a wide variety of services to research teams.

expertise

CERT has developed a strong expertise in DNA chips, high-throughput real-time PCR and "new generation" sequencing, including single cell sequencing. The following equipment is available to perform gene expression measurements:

Equipment for the qualification and quantification of nucleic acid samples

  • Spectrophotometry: Nanodrop 2000
  • Spectrofluorometry (QuBit 4.0 ThermoFisher Scientific) available on the CRCT platform
  • Microfluidic electrophoresis (Fragment Analyser) available on the CRCT platform

Transcriptomics equipment

  • Quantitative PCR
    • CFX96® ( 96 wells)-Biorad
    • StepOne (96 wells)-Applied
    • 2x LC480 (384 wells)-Roche
  • DNA microarrays
    • Affymetrix GeneAtlas Station
    • NanoString System
    • nCounter Analysis System. Uses molecular barcodes to detect and count hundreds of unique transcripts (without amplification).

The realization of NGS projects can be done with the CRCT platform.

The conditions of access are as follows :
Have contacted the Transcriptomics Manager Have received training on the instruments and techniques envisaged Have booked the instruments on the online booking schedule

We offer you advice and support to choose, adapt, and realize :

  • Construction of biomarkers, biosensors and fluorescent reporters,
  • genome editing to modify cell lines or small animals,
  • Validation of the constructs, using appropriate imaging, cell biology or biochemistry techniques.

CONTACT

emmanuelle arnaud

Emmanuelle Arnaud

Transcriptomics Engineer
odilemondesert

Odile Mondésert

Genomics Engineer

The platform's engineers evaluate the user's request and propose the most suitable solution for his care by :

  • providing advice in the development of experiments,
  • training users,
  • assisting users in the analysis and interpretation of their results,
  • accompanying users in the implementation of research and development projects, in the form of collaboration.

Physioxia & Hypoxia

Physioxia
& Hypoxia

Within tissues, the oxygen level is much lower than that of the atmosphere in which cells are conventionally grown (21% O2). In order to study cell metabolism in conditions close to those present in tissues, it is essential to grow them in an oxygen-controlled atmosphere. Cell oxygenation, at the heart of Inserm and Restore's research, has been awarded a prize in 2019.

expertise

This incubator mimics the physiological oxygenation conditions of cells or tissues throughout their culture, without fluctuation of the atmospheric environment. It allows the study of different populations of stem cells in conditions close to those encountered in vivo and to lead to new therapeutic leads. (complete article).

The X-Vivo apparatus (Biospherix) is an enclosure with a fully controlled oxygen atmosphere. It allows, thanks to its different modules, to realize simultaneously different cultures with different levels of oxygen (1%, 2%, 5%, …) according to the experimental demands and this throughout the culture:

  • 6 independent incubators with specific oxygen control

  • 2 independent working modules with specific oxygen control

  • 2 independent transfer airlocks

  • 1 module with a microscope

CONTACT

Métabolisme, adipocytes, métabolomique, fluxomique, traçage isotopique in vitro/in vivo, culture cellulaire en hypoxie, activité mitochondriale

Jeanson Yannick

Yannick Jeanson

PhD- Ingénieur Inserm

Vieillissement, fragilité, cancer, métabolisme redox, hypoxie, flexibilité métabolique, lipides, fibroblastes, cellules souches mésenchymateuses

ADER Isabelle

Isabelle Ader

PhD- Chargée de recherche Inserm

A researcher and an engineer, both experts, evaluate the user's request and :

  • Authorize access to the equipment after prior training
  • Advise for the design of experiments in hypoxia
  • Realize as a service your entire hypoxia experiment

Fluxomics

Fluxomics

In recent years, the study of metabolism has been at the heart of physiology and health issues. The applications are numerous and include the fundamental understanding of physiology and pathologies, the identification of new therapeutic targets and individualized post-therapeutic follow-up. The ability to understand metabolism is therefore a major challenge in the understanding of aging and in the fight against diseases such as cancer.

As part of a joint initiative between RESTORE, CRCT and the MetaToul platform, CERT houses an in vivo fluxomics facility that includes:

  • two modules allowing the realization of isotopic tracing experiments (stable isotopes: 13C, 15N, etc) in vivo or in vitro
  • the equipment necessary for the collection of samples and the extraction of metabolites
  • a culture chamber under controlled oxygen pressure to understand the effect of oxygen partial pressure variations on metabolism
  • a respirometry tray to measure oxygen consumption flows

Metabolome, lipidome and isotope labeling analyses are performed on the MetaToul platform, which has an instrumental park (mass spectrometry, NMR) and unique expertise in this field in France. The whole set-up and the associated expertise represent also a unique configuration in France for in-depth studies of metabolic dynamics at different scales (from the cell to the whole organism).

expertise

The study of metabolic flows within an organism (intra or inter-tissue) is a major issue in order to study metabolic deregulations observed during aging or during the development of pathologies such as metabolic diseases or cancer.

in vivo Fluxomics module

It contains the infrastructure and equipment necessary to perform a complete in vivo isotope tracing study on mice:

  • Establishment using animals for scientific purposes approved by the DDPP
  • Gas anaesthesia tables for the catheterization of mice
  • Device for perfusion of 13C-labeled substrates (or other stable isotopes) on vigorous mice (syringe pumps, portals, swivels)
  • Tissue collection and preparation, sample preparation for analysis.

A large body of literature demonstrates that metabolism impacts cell fate and orientation. The study of metabolic fluxes at the cellular level is fundamental to better understand the biology of the cell and to predict its loss of function during aging or the development of metabolic diseases or cancer.

in vitro Fluxomic module

This module has the necessary equipment to carry out an in vitro isotope tracing study in its entirety: cell culture under controlled oxygen atmosphere (see culture under controlled oxygen atmosphere), labelling experiments with 13C-labelled metabolites (or other stable isotopes), sampling and extraction of internal (endo-metabolome) or external (exo-metabolome) metabolites:

  • Fully oxygen controlled atmosphere chamber (X-Vivo, Biospherix)
  • Various equipment for endo- and exo-metabolome extraction
  • Quenching system (immediate blocking) of the in vitro metabolism

The mitochondrion is the major site of ATP production within the cell and plays a key role in cellular redox metabolism. Mitochondrial activity can be modified during the development of pathologies such as metabolic diseases, cancer or during aging. This mitochondrial activity can be evaluated by measuring the oxygen consumption with an oxygraph.

Oxygraphs

The Oroboros type oxygraphs present on the CERT platform are very sensitive instruments due to their hermetic tanks and Clark probes. They allow the study of mitochondrial activity on tissues (homogenates, permeabilized), cells and isolated mitochondria using different substrates and drugs. The Oroboros type oxygenographs are not limited in the number of injections per experiment which allows a more precise study of mitochondrial activity:

  • 2 Oroboros oxygraphs

  • 2 independent tanks per oxygenograph

CONTACT

Métabolisme, adipocytes, métabolomique, fluxomique, traçage isotopique in vitro/in vivo, culture cellulaire en hypoxie, activité mitochondriale

Jeanson Yannick

Yannick Jeanson

The platform engineer evaluates the user's request and proposes the most suitable solution for his needs. He :

  • provides advice in the development of experiments,
  • trains the users,
  • assists users in the analysis and interpretation of their results,
  • assists users in the implementation of research and development projects, in the form of collaboration.

Models of aging

Nothobroanchius

Models of aging

The regeneration of tissues and organs represents a major cognitive and medical challenge. The objective of our institute is to understand some of the underlying mechanisms to improve/restore regenerative capacities in adults.

The institute offers expertise in several models of aging, particularly in fish such as Brachyodanio rerio and Nothobroanchius furzeri.

These species were chosen mainly for their homology with the phenomena studied in humans (physiological, clinical…), for the technical tools that can be implemented (genetic tools allowing for example access to mutated lines, physical tools such as imaging, surgical tools allowing the development of models and the use of biomaterials) as well as for the bibliographic data related to these species.

expertises

L’étude des flux métaboliques au sein d’un organisme (intra ou inter-tissulaire) est un enjeu majeur afin d’étudier les dérégulations métaboliques observées au cours du vieillissement ou lors de la mise en place de pathologies telles que les maladies métaboliques ou le cancer.

Module de fluxomique in vivo

Il contient les infrastructures et les appareils nécessaires pour réaliser l’intégralité d’une étude de traçage isotopique in vivo sur souris:

  • Etablissement Utilisateur d’animaux à des fins scientifiques agréé par la DDPP
  • Tables d’anesthésie gazeuse pour la cathétérisation des souris
  • Dispositif de perfusion de substrats marqués au 13C (ou autres isotopes stables) sur souris vigiles (pousse-seringues, portiques, swivels)
  • Prélèvement et préparation des tissus, préparation des échantillons pour l’analyse.

Une littérature fournie démontre que le métabolisme impacte le devenir et l’orientation cellulaire. L’étude des flux métaboliques au niveau cellulaire est fondamentale pour mieux comprendre la biologie de la cellule et prédire ses pertes de fonction au cours du vieillissement ou de la mise en place des maladies métaboliques ou du cancer.

Module de fluxomique in vitro

Il dispose des équipements nécessaires à la réalisation d’une étude de traçage isotopique in vitro dans sa globalité : culture cellulaire sous atmosphère contrôlée en oxygène (cf culture sous atmosphère contrôlée en oxygène), expériences de marquage avec des métabolites marqués au 13C (ou autres isotopes stables), échantillonnage et extraction des métabolites internes (endo-métabolome) ou externes (exo-métabolome):

  • Enceinte à une atmosphère entièrement contrôlée en oxygène (X-Vivo, Biospherix)
  • Divers équipements pour l’extraction de l’endo- et de l’exo-métabolome
  • Système de quenching (blocage immédiat) du métabolisme in vitro

La mitochondrie est le site majeur de la production d’ATP au sein de la cellule et joue un rôle primordial dans le métabolisme redox cellulaire. L’activité mitochondriale peut être modifiée lors de la mise en place de pathologies telles que les maladies métaboliques, le cancer ou encore lors du vieillissement. Cette activité mitochondriale peut être évaluée grâce à la mesure de la consommation en oxygène à l’aide d’un oxygraphe.

Oxygraphes

Les oxygraphes de type Oroboros présents sur le CERT sont des instruments très sensibles grâce à leurs cuves hermétiques et leurs sondes de Clark. Ils permettent d’étudier l’activité mitochondriale sur tissus (homogénats, perméabilisés), cellules et mitochondries isolées grâce à l’utilisation de différents substrats et drogues. Les oxygraphes de type Oroboros ne sont pas limités en nombre d’injection par expérience ce qui permet une étude plus précise de l’activité mitochondriale :

  • 2 oxygraphes Oroboros

  • 2 cuves indépendantes par oxygraphes

CONTACT

Foie, tissu adipeux, macrophages, métabolisme, inflammation, vieillissement

user

Jean-Philippe Pradere

L’ingénieur de la plateforme évalue la demande de l’utilisateur et propose la solution la plus adaptée à ses besoins. Il :

  • assure le conseil dans l’élaboration des manipulations,
  • forme les utilisateurs,
  • assiste les utilisateurs dans l’analyse et l’interprétation de leurs résultats,
  • accompagne les utilisateurs dans la mise en place de projet de recherche et développement, sous forme de collaboration.