Manager: David Jarrossay, Diego Morone

This imaging facility is central to most of the IRB research projects. The IRB has a state-of-the-art flow cytometry facility that is run by David Jarrossay  who takes care of the cell sorting, maintenance of the equipment, instruction and advice to the new operators, in addition to performing his own research. Diego Morone has a specific experience in confocal microscopy and high content cellular analysis and supervises all the microscopes present at the IRB.

Flow cytometry
The Flow Cytometry lab provides investigators with equipment and support for cell sorting (separation), acquisition, and analysis of flow cytometric data with a variety of state-of-art multicolor flow cytometry instruments.
Cell sorting is performed on a FACSAria III equipped with four lasers (488, 561, 640 and 405 nm excitation wavelengths) and 15 fluorescence channels detection. It can perform high speed sorting (up to 20, 000 events/sec) with high-purity (up to 99%).

Benchtop analyzers
The Flow cytometry lab is equipped with an advanced benchtop analyzer BD Fortessa equipped with four lasers (488, 561, 640 and 405 nm excitation wavelengths) and 16 fluorescence channels, one FACSCantoII (488, 640 and 405 nm lasers-eight colors), a FACSCanto I (488 and 640 nm lasers-six colors). The lab also acquired a SP6800, a Spectral Cell Analyzer from Sony (488 and 640 nm lasers excitation wavelengths with 405 nm laser to come) equipped with 32 channel PMT collecting spectra of all emitted lights with no need for conventional bandpass filters.

High throughput screening
Beside both FACSCanto equipped with HTS for acquisition of 96 and 384 wells plates, the lab has also been equipped with an Intellicyt (488 and 633 nm excitation-four colors) with automated platform (up to 45 plates per run) allowing high throughput screening for 96 or 384 well plates format.
The flow lab offers efficient support and high quality instruments. The staff provides cell sorting on BD FACSAria III, individual training on bench top analysers, maintenance of all instruments and assistance with experimental design, data analysis and troubleshooting.

Imaging
The Microscopy Facility at the IRB provides state-of-the-art imaging equipment and dedicated expertise for the most demanding research projects:

• Confocal microscope. Leica TCS SP5, fully equipped with 5 lasers, 4 new-generation hybrid detectors (HyD Leica technology), fast z piezoelectric stage for fast 3D live cell imaging, resonant scanner and aberration-corrected objectives for high-resolution imaging such as 20X multi-immersion objective or 100X 1.4 Oil objective.

• High-content screening system. Molecular Devices ImageXpress Micro 4, wide-field automated microscope for up to 5 fluorescence channels and transmitted light. The microscope is combined with a robot for automated acquisition and analysis of up to 45 plates. Software allows a very simple acquisition setup and provides modules for segmentation and analysis.

• Wide-field microscopes
– Nikon Eclipse E800 upright microscope;
– Nikon Eclipse TE300 inverted microscope, with incubator for live-cell experiments and Eppendorf FemtoJetmicroinjector;
– Zeiss Axiovert 200 inverted microscope, equipped with UV-corrected optics and TILLvisION software for ratiometric calcium measurement experiments.

• Multi-Photon Excitation Microscopy system.
LaVision BioTec TriM Scope combines both an upright and inverted microscope, equipped with two tunable pulsed NIR lasers and OPO for multi-colour simultaneous imaging on up to 5 channels.

With these systems we are able to perform most of the procedures for cell and tissue imaging, such as FRAP, FLIP, photoconversion or FRET and 3D reconstruction on both unclarified and clarified tissues.
We also offer support for sample preparation, image analysis, deconvolution and 3D reconstruction, thanks to a wide range of open software such as ImageJ / FIJI, R, CellProfiler, GNU Octave, as well as commercial ones, such as MATLAB (MathWorks), MetaMorph (Molecular Devices) and Imaris (Bitplane).\