Light-Sheet Fluorescence Microscopy
Local Service Provider

Light-Sheet Fluorescence Microscopy (LSFM) of cleared brain samples offers the unique opportunity of exploring brain complexity on large scales while keeping cellular resolution. LENS has gathered long-standing experience in the whole LSFM pipeline, comprising sample preparation (clearing and staining), imaging using custom-designed optical setups, and image post-processing (volumetric stitching, compression, feature segmentation).

Part of the proposed EBRAINS Ecosystem of Local Service Providers, the LSFM LSP offers access Light-Sheet Fluorescence Microscopy setups, Two-Photon Fluorescence Microscopy setups and a Spinning Disk Confocal. Within the LSP, LENS can provide clearing and staining of the samples, imaging, and data processing (deskewing, stitching, cell localization or 3D fiber analysis) or can host external scientists that wish to use the microscopes or acquire technical skills in light-sheet microscopy. Specific projects need to be discussed beforehand to ensure optimal planning of resource usage and the organisation of specific training if needed.

Data obtained with LSFM can be connected with that obtained from other physical facility in order to obtain multimodal analysis of the same tissue. In particular, samples can be acquired both with MRI and 3D-PLI before LSFM measurement, and can undergo SIB-SEM analysis after it. In addition, the 3D point clouds of the cells obtained from the analysis of the LSFM reconstruction can be used to perform modeling in the Cerebellar, Hippocampus or Basal Ganglia Modelling LSPs, who are also part of the EBRAINS LPS network.

• Two-Photon: A multimodal optical platform integrates two-photon excitation microscopy, spectrally-resolved fluorescence lifetime imaging microscopy, Raman micro-spectroscopy and diffuse reflectance micro-spectroscopy into the same device. The combination of these different techniques greatly enhances the morpho-functional investigation of biological tissues (either bulky samples or thin sections) in a fast and label-free modality. There are no particular restrictions on sample size, although the penetration depth of these optical techniques is limited to few hundreds microns.
• Two Light-Sheet Fluorescent Microscopy setups capable of high-speed and high-resolution imaging with up to four-color acquisition. One system can accommodate large bulky samples such as a whole mouse brain, while the other microscope is designed for thin but large tissue slices (several cm in lateral size (max 10cm), 0.5mm to 1mm in thickness).
• Spinning Disk Confocal (Nikon Crest X-Light V3) integrated with Nikon’s microscope platforms and optics, capable of wide array of outputs. It has a large field homogeneous illumination (up to 25 mm FOV) allowing for quantitative imaging over large FOVs and high-speed imaging on large samples. It also features dual camera imaging at 25 mm FOV. Different camera formats can be chosen for the two ports, making it very flexible.

Local Service Providers (LSPs) are institutions, facilities, competence centers and groups that possess unique capabilities for top of the line neuroscience, including but not limited to databases, tools and softwares, advanced technologies, expertise and personnell. They are part of a proposed ecosystem of collaborative institution that are part of EBRAINS.

The LSFM Local Service Provider is part of the EBRAINS Italian National Node, centred on the Multimodal, Multiscale, and Multi-model analysis of the spatiotemporal organization of the brain (3M approach). Coordinated by the National Research Council, it involves multiple research centers across the country