Facilities and Equipment

The CEMIPAI platform houses a 360 m2 enclosed laboratory facility of biosafety level 3 (BSL3). All procedures involving infectious material are done within dedicated biological safety cabinets installed in a total of 8 individual boxes  

 

Biosafety Cabinets

6 boxes of the BSL3 facilities are dedicated to either viruses or bacterias and contain state-of-the-art equipment, including: 

Flow Cytometry

A box equipped with a NovoCyte Flow Cytometer 2100Y installed in a Biological Safety Cabinet. 

 

  • 2 lasers (blue 488nm and yellow 561nm), which can be upgraded with a red or violet laser.
  • 10 fluorescence parameters
  • Absolute counting, without addition of counting beads
  • Sample tube/plate holder (24 tube rack or 24-48-96 well plates)
  • NovoExpress Software is enable to analyze cell cycle with Watson’s method (Cytometry 2001 ;43:55-68)

 

High Throughput Screening

Our BSL3 is equipped with a workstation specifically designed for automated drug screening on cell-based assays

  • A Zephyr multichannel liquid handler with 96-and 384-well capabilities for performing accurately and efficiently plate preparation, replication or reformatting, serial dilutions and cell based assays.
  • A Biotek plate washer.
  • An Envision Multilabel reader (Perkin Elmer). This fast and effective reader can accommodate a wide range of mirror modules and filters for a variety of detection approaches based on fluorescence, luminescence and absorbance.
  • A Twister II Microplate Handler with multiple instrument capability, integral plate and tip storage capacity.
  • A microplate incubator LiCONiC STX44 (2 racks for a capacity of 44 microplates)

The workstation is supported by iLinkPro, a robust and easy to use control software that connects all the instruments to set up a fully automated assay system.

Screening BSL3 Robot CEMIPAI

Electron Microscopy Facilities

Depending on your question and sample we offer a variety of different preparation techniques to ensure optimal preservation of your specimen:

  • Preparation for transmission electron microscopy (TEM) to study surface and pathogen structures
  • Conventional chemical fixation for tissue samples and cell monolayers.
  • Immuno-labeling to locate structures of interest.
  • Negative Staining for rapid visualization of macromolecules, proteins, viruses, organelles or bacteria.

A) TEM micrographs of ZIKA virus-infected primary fibroblasts. Membrane vesicles with sizes between 70 and 100 nm, observed in intimate association with the endoplasmic reticulum, are indicated by a white arrow. Black arrows indicate the presence of spherical capsids detected in intracellular vacuoles or docked to intracellular membranes. From Hamel et al., J. Virol. 2015 Sep 89(17) 8880-96 

B) TEM micrographs of wt-HIV particles

C) TEM analysis of influenza A(H1N1) Virus-Like Particle production in HEK 293T cells. From Kerviel et al., PLoS One. 2016 Nov 4;11(11):e0165421. doi: 10.1371/journal.pone.0165421

D) U2OS cells transiently transfected with Coxiella Burnetii vacuolar protein B (CvpB)-HA, fixed and processed for immuno-EM with the HA tag labeled using Nanogold particles. CvpB could be detected in clusters of small vesicles, enlarged vesicles or plasma membrane protrusions. From Martinez et al., Proc Natl Acad Sci U S A. 2016 Jun 7;113(23):E3260-9. doi: 10.1073/pnas.1522811113

Super Resolution Microscopy TIRF-PALM (Photonic imaging)

PALM microscope in TIRF mode for super-resolution and live cell imaging

Installed in the BSL3 facilities, our Nikon Ti-E microscope is equipped with 3 different objectives (20x long WD (1.2 mm), oil immersions 40x and 100x corrected for super resolution imaging) and 5 different filter cubes, 3 classical for blue (Dapi), green (FITC) and red (mCherry) imaging, 2 optimized for super resolution (STORM or PALM w/mEos2). The EM-CCD detector allows fast acquisition in low fluorescence levels. 3 different lasers, namely 405, 488 and 561 nm are fiber coupled to a TIRF module allowing either TIRF excitation or wide-field excitation. A perfect focus system is also available to avoid any z-drift during long time image acquisition.

  • Inverted microscope NIKON Ti-E for live-cell data acquisition
  • Epi-FI LED illuminator
  • Diodes Laser: 405nm, 488nm, 561nm, 641nm / High N.A. objectives for live cell imaging : 20x, 40x, 100x
  • Detector: EMCCD Camera IXON 897 (512Ă—512 chip, pixel 16µm)
  • Multi-channel (multi-color) image acquisition
  • TIRF arm
  • Motorized stage XYZ with autofocus system PFS for multi-dimensional time-lapse imaging
  • Z-series image acquisition
  • Software: NIS

The microscope is inserted into an incubation room to maintain biological viability as long as needed. Acquisition and interfacing software are provided by Nikon.

High-Speed Atomic Force Microscopy on living cells

 

Since March 2018, our BSL3 facility is equipped with the latest generation of atomic force microscope (AFM) designed for real time imaging of cells and class-3 pathogens.  Our NanoWizard4 BioScience from JPK Instruments combines nano-scame resolution and fast scanning, integrated with an optical microscope. It is designed to provide high resolution AFM coupled with fluorescence optical imaging  during long term experiments on samples ranging from single molecules, viruses, bacteria to living eukaryotic cells and tissues. This set-up in a L3 laboratory is unique in Europe.

Features

 

  • JPK Nanowizard 4 Biosciences designed and modified for BSL3 environment and live cell imaging of pathogens (bacteria, virus) and infected cells
  • Integration with a NIKON Ti inverted microscope for fluorescence imaging during AFM acquisition
  • Fast scanning of up to 100 lines/sec and time per frame of up to 3.6 sec for tracking dynamic processes
  • Nanometer-scale resolution with a large scan field of 100 Ă— 100 Ă— 15 µm3
  • High-resolution quantitative imaging by QI™ quantitative imaging mode
  • DirectOverlay™ software for direct correlation of AFM and optical data
  • Thermoregulated liquid cell for live cell imaging
  • Epi-FI LED illuminator for fluorescence imaging at 488nm and 561nm