Photonic Microscopy

The Cellular Imaging Facility offers imaging techniques from the sub-cellular level to the scale of the organism, as well as flow cytometry. 6 engineers (2 IR-CNRS, 1IR-SU, 2 IE-CNRS and 1IE-INSERM, 20%) ensure the maintenance and quality control of the equipment, the training and support of users as well as the technological and methodological development and scientific animation.

The Cellular Imaging facility provides access to a large variety of optical instruments (several confocal microscopes, including 2 coupled to a Becker & Hickl FLIM module, spinning disc/ILAS II microscope, TIRF/video microscope, light sheet microscope, multiphoton microscope, macro-apotome, multilaser analyzers).

The facility's investments were possible thanks to strong support from the CNRS, Sorbonne-University, the Île-de-France Region and GIS-IBISA, as well as from multiple teams and laboratories.

The facility proposes new imaging and flow cytometry methods (F-techniques, FLIM, 3D imaging, spectral imaging, life imaging, multicolor analysis) in collaboration with research groups, and puts its expertise at the service of the entire scientific community.

Equipment

The offer, accessible by online reservation, is aimed at all research players (IBPS, P6 campus, public research organizations, private companies).

- ZEISS 980 FAST-Airyscan II upright microscope coupled to FLIM Becker&Hickl

- ZEISS 980 FAST AIryscan II inverted coupled to FLIM Becker&Hickl

- Upright confocal microscope Leica TCS SP5 AOBS 

- Inverted confocal microscope/resonant scanner Leica TCS SP5 AOBS 

- Upright Biphoton/confocal microscope / resonant scanner Leica TCS SP8 MPII

- Phaseview Alpha 3 Lightsheet Microscope

- SPINNING DISK /ILAS II upright microscope

- NIKON TIRF/inverted Video Microscope

- Macro-apotome

- VYB Analyzer

- MACSQuant Analyzer

- 3 data processing and analysis stations with ImageJ, Huygens, Metamorph, Volocity, FlowJo, Venturi-One

Contact and services

To contact please send an email to :

imagerie.ibps@listes.upmc.fr

Offers may include conception monitoring, follow-up, realization, analysis and result analysis of possible experimental strategies on our equipment.

Type of offers :

  • Observation sessions for autonomous users : Autonomous use of equipment, attribution of a user account, data availability.
  • Engineer-assisted observation session : Image acquisition monitored by an engineer, attribution of a user account, data availability.
  • Long-term collaborative project

Expertise

- Assistance with sample preparation (clarification, immunolabeling advice, etc.)

- Three-dimensional imaging from the cell to the whole organism

- Life Imaging 

- Photomanipulation (FRAP, Photoconversion, Photoablation)

- Spectral imaging

- Fluorescence Lifetime Imaging

- Image restoration and analysis: Co-localization analysis, deconvolution (wide-field,    confocal, spinning disc, multiphoton), 3D reconstruction...

RESEARCH & DEVELOPMENT

We are currently following 4 axes of R&D:

1) Preparation of complex biological samples 

2) Optimization of the 3D-resolution , Confocal Super-resolution 

3) Design of 3D spatial analysis tools 

4) Optical instrumentation for fast and 3D imaging

R&D sample preparation and data processing

Development of clarification protocols for complex biological samples:

A biological sample is composed of different molecules with refractive indices varying between 1.33 (water) and 1.56 (proteins). The sample is therefore opaque because the light used for imaging is strongly scattered, making deep imaging very difficult. Therefore, complex biological samples must be made transparent to allow volume imaging. Over the past ten years, we have observed a revolution in volumetric imaging of complex samples thanks to the clarification technique.

Clarification makes it possible to observe internal structures present in opaque and thick biological samples, called complex (tissues, organs or small embryos). It makes volumetric optical observation accessible. The principle of clarification consists in homogenizing the differences in refractive index, in order to obtain transparent biological specimens.

Clarification methods must be adapted to each biological sample, which explains the multitude of protocols. The method used will depend on the nature of the sample, its size, the rate of auto-fluorescence, its complexity and the desired fluorescent labeling.

We are developing, in collaboration with research teams, new low-toxic clarification protocols, allowing the imaging of tissues, small embryos and spheroids of multiple biological organisms (mice, chicken, fruit fly, xenopus, axolotl, grass snake, sea dogfish, etc.).

In addition, we offer training to guide the user in the multitude of protocols that exist, starting from a theoretical introduction to the range of existing techniques and observation methods ending with the prerequisites for a suitable data analysis.

R&D confocal super-resolution

Optimization of 3D resolution: 

We have worked to implement confocal super-resolution by optimizing sample preparation (Fouquet et al., 2015), followed by developing a workflow of optimisation of acquisition parameters and systematic deconvolution of images in order to substantially double the lateral and axial resolution in confocal microscopy (Lam et al., 2017)

We arena studying the improvement of axial resolution in thick fixed samples (mouse brain, zebrafish, etc.) immunolabeled or carrying fluorescent proteins in complex biological samples and in depth. 

We are currently developing confocal super-resolution thanks to the acquisition of two ZEISS 980 Fast Airyscan II and the deconvolution of data from this type of imaging. We are also studying the artifacts of this kind of image processing.

R&D Spatial analysis in 3D

Development of spatial analysis tools: 

The facility team has proven expertise in the analysis of cellular events such as co-localization (Bolte and Cordelières, 2006, Cordelières and Bolte, 2014).

In the past, we used “JaCoP”, a plugin for ImageJ. This analysis tool is restricted because it is based on intensities. This is the reason why we have recently developed a more advanced tool for the analysis of cellular structure associations based on object detection and 3D spatial relationship analysis. This automated tool is called DiAna (Distance Analysis, Gilles et al., 2017) and was designed in collaboration with Dr. T Boudier (Centuri, Marseille). 

A new ImageJ plugin project aims now to facilitate the study of dendritic spines. This work is done in collaboration with N. Heck and T. Boudier and takes advantage of two recognized ImageJ plugins, 3D ImageJ Suite and SNT. Its purpose is to study the morphological characteristics of spines based on a semi-automatic approach and by exploiting their shapes in 3D for more precise results.

R&D Instrumentation

The facility has a strong partnership with the Jean-Perrin Laboratory (UMR CNRS/SU 8237), a constituent unit of the IBPS. This laboratory mainly brings together physicists interested in the challenges of biology. He has recognized expertise in optical instrumentation for biology, and was notably a pioneer in the development of light sheet microscopy for functional imaging.

As part of the attachment of the Jean Perrin laboratory to the IBPS, the IBPS imaging facility is thus a stakeholder in several federative instrumental projects associating several sites of the LUMIC network (Brain and Spinal Institute, ICM and L Institute of Vision, IdV), which aim to set up new innovative imaging systems (high spatial and temporal resolution light sheet microscopy). These have recently been supported by IDF and GIS-IBISA funding

It is Thomas Panier (IR-SU) who is responsible for the instrumentation projects.

Online reservation

Partners

The IBPS imaging facility is part of the national networks GDR IMABIO and rt-mfm (France LAM, Organization des Assises RTmfm, Communication Manager). In addition, the facility coordinates the imaging and cytometry network of Sorbonne- University LUMIC (Susanne Bolte, operational manager).

Training

The imaging platform is a strong player in training at local, national and international level: 

CNRS Corporate Training: Clarification for three-dimensional imaging of complex biological objects, annual 

CNRS-MiFoBio: 7 Workshops on confocal super-resolution, clarification and spatial analysis. 

CNRS IFSEM-training: Processing and analysis of microscopy images using ImageJ software, Level I: Initiation, annual 

INSERM workshops: 

Workshop 241: Live imaging of morphogenesis in 4D: probes, microscopy techniques and Quantification 

Workshop 264: Tissue clearing, sample preparation, 3D imaging and data analysis

Operational Manager

TEAM

Scientific support staff

Other (volunteers, sabbatical...)

NEWS

Jean-François Gilles was awarded the CNRS Cristal Medal for his development of a plugin, Distance analysis (DiAna), for the ImageJ software dedicated to image processing and analysis for biomedical research.

DiAna, which can be used with images informed on the basis of metadata, and which also integrates options for the segmentation of objects, is a response to a strong need of the international scientific community since it makes it possible to calculate the spatial colocalization of 3D objects and measure the distance between them.

New Materials

Two brandnew ZEISS 980 Airyscan II confocals coupled to fluorescence lifetime imaging have arrived at the facility. 

You want to know more? Contact us at:

imagerie.ibps@listes.upmc.fr

Acknowledgement of imaging facility in your publications

It is COMPULSORY to acknowledge the imaging facility in your publications if you present data that has been produced with the help of facility equipment or personnel. If the participation of an engineer is décisive you should consider co-authorship.

The development of the imagine facility is possible du to funding by several sources. In order to acknowledge the facility and the personnel the following phrase must be used every time you publish work including equipment and/or the help of engineers:

Image acquisition and/or image analysis were performed at the IBPS Imaging Facility the authors greatly acknowledge ________ of the IBPS Imaging Facility. The IBPS Imaging facility is supported by Region-Île-de-France, Sorbonne-University and CNRS.

Thank you to indicate accepted papers to the staff, wee need them for our record to the sponsors/donors.

Collaborations

Dr. Thomas BOUDIER, CENTURI mutli-engineering platform, Marseile

Dr. Michel GHO, Laboratoire de Biologie du Développement, IBPS, Paris

Dr. Nicolas HECK, Neurobiologie Paris-Seine, IBPS, Paris

Dr. Eglantine HEUDE, Laboratoire Physiologie moléculaire et Adaptation, MNHN, Paris

Dr. Katja WASSMANN, Laboratoire de Biologie du développement, IBPS, Paris

BIOM, Observatoire Océanologique de Banyuls-sur-mer

PHASEVIEW