SPAIN

Barcelona Live and Intravital Node

The Barcelona Live and Intravital - Advanced Light Microscopy Node (BLivIN) provides expert support for biomedical projects requiring functional live-cell and in vivo imaging. BLivIN offers expertise in a wide range of advanced and innovative technologies, including widefield fluorescence and transmitted optical microscopy, laser scanning and spinning disk confocal microscopy, two-photon excitation (2PE), second harmonic generation (SHG), Total Internal Reflection Fluorescence (TIRF) and image scanning microscopy (ISM), time-correlated single photon counting, lifetime imaging (or FLIM) and laser photomanipulation. Additionally, the Node supports functional imaging technologies with robust capabilities in image processing, analysis, and custom-developed solutions.

To enhance the success of visitor projects, BLivIN provides tailored planning and comprehensive support throughout all project stages, from experimental design and sample preparation to acquisition, image processing and analysis. Visitors also gain access to complementary resources such as animal facilities, wet labs, cell culture capabilities, and non-invasive bioluminescence imaging.

Specialties and expertise of the Node

Our node provides scientific and technological expertise to extract functional, dynamic, and high-content information from living biological systems through advanced bioimaging applications. Our areas of expertise include:

• Functional imaging: Techniques based on photobleaching and photoactivation, Fluorescent Resonance Energy Transfer, FRET ratio imaging, Lifetime imaging (FLIM) based on Time-Correlated Single Photon Counting and FLIM/FRET imaging.
• High-content and high throughput automated live-cell imaging: Long-time lapse, multi-position imaging (mosaics, multi-well plates), live-cell high-content screening, smart screening and rare event detection. Applications include cell tracking, wound healing, autophagy, cell viability, mitochondrial health, protein synthesis, transcription factor activation, cell cycle, mitotic index, cytoskeletal rearrangement, apoptosis (TUNEL), cell proliferation, endocytosis and oxidative stress.
• Two-Photon in vivo imaging: Deep tissue imaging of organs (brain, skin, liver, kidney, muscle, lymph nodes, eye) in rodents and small animals (Xenopus, Drosophila, zebrafish). Imaging at molecular, subcellular, and cellular levels using transgenic fluorescent animals or intravenous vital fluorescent contrast agents.
• Non-linear applications: Functional live-cell imaging, photo-stimulation assays (optogenetics, uncaging, photothermal and photochemical studies), SHG for collagen, myosin and starch imaging and third harmonic generation (THG) for lipids studies. Imaging of UV fluorochromes such as Laurdan and Filipin.
• Fast optical sectioning: Techniques for living samples including image-scanning microscopy and spinning-disk microscopy.
• Laser nanosurgery: Ablation in living cells and tissues using pulsed laser irradiation, combined with spinning disk or widefield microscopy. Applications include DNA damage kinetics, wound healing, subcellular damage and intra- and inter-cellular (mechanical) tension release, …
• Bioimage analysis: Our team of four BioImage analysts develops advanced workflows to integrate the processing, analysis, quantification and visualisation and interpretation of Bioimage data. We offer training in commercial and open-source software tools. When these tools are insufficient to extract results, we develop custom workflows that combine various packages and algorithms. Special tools for in vivo imaging tracking and custom deconvolution are also available.
• Complementary imaging techniques and services: Multimodal or correlative imaging capabilities combined with live/ in vivo imaging applications, such as correlative light and electron microscopy (CLEM), super resolution imaging , light sheet- based imaging and histological sectioning.

Offered Technologies

• Deconvolution widefield microscopy (DWM)
• Laser scanning confocal microscopy (LSCM/CLSM)
• Spinning disk confocal microscopy (SDCM)
• Total internal reflection fluorescence microscopy (TIRF)
• Two-photon microscopy (2P)
• Image Scanning Microscopy (ISM)
• Second/Third Harmonic Generation
• High throughput microscopy/high content screening (HTM/HCS)
• Fluorescence Resonance Energy Transfer (FRET)
• Fluorescence Recovery after Photobleaching (FRAP)
• Fluorescence Lifetime Imaging (FLIM)
• Fluorescence (cross)-correlation spectroscopy (FCS/FCCS)
• Intravital Microscopy (IVM)
• Microdissection *
• Photomanipulation (Pmnip)
• Intravital microscopy (IVM) - Med
• Image Analysis-bio *

Additional Services offered by the Node

• Instruments

• Methodological setup (e.g. design of study protocol and standard operation procedures)

• Technical assistance to run instrument

• Training in infrastructure use

• Training in techniques for optical clearing of biological samples

• Probe preparation

• Biological material storage and processing

• Animal facilities (Rodents, Zebrafish, Xenopus laevis, Xenopus tropicalis, Drosophila melanogaster)

• Wet lab space

• Data processing and analysis

• Server space

• Training workstations

• Training seminar rooms

• Housing facilities

• Regulatory affairs management service

Instrument highlights

• Laser nanosurgery (custom system) based on subnanosecond pulsed laser yielding submicron damage control.

• Lifetime imaging through visible pulsed lasers (including white laser), FLIM analysis in the time domain and phasor plot analysis.

• Image scanning microscopy (e.g. Airyscan) provides super resolution and/or fast imaging capabilities for live samples.

• Fast spinning disk with parallel cameras for synchronous multichannel imaging

• High-content imaging and analysis of multiwell plates includes plates robot-loading and in situ incubation for long term (days, weeks) time lapse imaging. Primary and secondary screenings.

• Multiphoton microscope with motorized bridge-type upright configuration, translation over sample, high-speed resonant and conventional scanners, three high-sensitivity GaAsP detectors, lenses with high working distance and numerical aperture, corrected for infrared imaging, dual tunable pulsed laser (100fs, 80MHz) with a wavelength range of 660-1320nm and a fixed line at 1040nm.

Contact details

Maria Calvo- University of Barcelona
mariacalvo@ub.edu

Julien Colombelli- Institute for Research in Biomedicine- IRB Barcelona
julien.colombelli@irbbarcelona.org

Nadia Halidi- CRG - Centre for Genomic Regulation
nadia.halidi@crg.eu