SMALL ANIMAL MULTIMODAL€¦ · Imaging Service offers the Carestream In-Vivo Xtreme animal imaging system, which allows up to five mice to be visualized simultaneously using fluorescent

SMALL ANIMAL MULTIMODAL IMAGING SERVICE Magnetic Resonance Imaging (MRI) Fluorescent and Bioluminescent

Optical Imaging Computed Tomography (CT)

MAGNETIC RESONANCE SERVICE High Resolution Liquid NMR High Resolution Magic Angle

Spinning (HR-MAS) NMR Time Domain NMR (TD-NMR)

ADVANCED FLUORESCENCE SERVICE Confocal Microscopy Multiphoton Microscopy Super-resolution Microscopy Flow Cytometry / Cell Sorter High Content Screening

ELECTRON MICROSCOPY SERVICE Transmission Electron Microscopy (TEM) Cryo-TEM Scanning Electron Microscopy (SEM) Environmental SEM (ESEM)

Bionand’s Nanoimaging Unit brings together a wide range of cutting-edge technologies, including high-field magnetic resonance imaging and spectroscopy, micro-CT, intra-vital optical imaging, electron microscopy, advanced optical microscopy, cell sorting and high content screening. The unit has been conceived to support nanotechnology-based biomedical research at Bionand and to make these advanced services available to the widest possible range of external users from other research institutes, universities, hospitals and companies. The Unit is divided into four services, each with a dedicated specialist.

Small animal (pre-clinical) imaging is a vital tool in biomedical research that allows biological processes and potential medicines to be studied in animal models using non-invasive methods. The service offers some of the most relevant pre-clinical imaging modalities, namely, MRI, in vivo Bioluminiscence and Fluorescence Imaging, and micro-CT. These techniques are essential for preclinical evaluation and characterization of new diagnostic and therapeutic agents. The combined use of these technologies and new highly specific imaging probes are the basis of molecular imaging.

SMALL ANIMAL MULTIMODAL IMAGING SERVICE

Contact Dra. María L. García Martín Head Nano-Imagen Unit Phone +34 952 367 636 Ext. 638 E-mail [email protected]

Small Animal Multimodal Imaging Service

MAGNETIC RESONANCE IMAGING (MRI)

Resonance Imaging (MRI) is a Highly versatile technique that can Provide not only excellent morphological data, but also functional And Physiological information as well. Furthermore, important Recent Advances in the development of highly sensitive contrast Agents Using Nanotechnology have brought MRI into the molecular Imaging arena. Thus, small animal MRI scanners have become a key tool in the development of magnetic Nanoparticles for medical diagnosi and therapy. Equipment Bruker BioSpec 9.4T/20 animal MRI system Specifications

Magnetic field strength of 9.4T 20 cm bore diameter, suitable for mice and rats Automated positioning system (AutoPac™) High performance actively shielded

gradient system with integrated shims RF coils for rats and mice

Applications

Anatomical Studies: tumors, inflammation, Degenerative diseases, etc.

Functional Studies: brain perfusion, vascular permeability, in vivo metabolic profiling, etc.

Pharmocokinetics and Biodistribution of new Contrast Agents (or any drug magnetically labeled)

Monitoring of Drug Delivery and Targeting Follow up of Therapy Response

Small Animal Multimodal Imaging Service

FLUORESCENT AND BIOLUMINESCENT OPTICAL IMAGING

Contact Dr. John Robert Pearson Advanced Fluorescence Specialist Phone +34 952 367 634 Ext. 634 E-mail [email protected]

The use of fluorescent and bioluminescent techniques for small animal-based studies into cancer, arthritis and heart disease have grown tremendously in recent years. The Bionand Multimodal Imaging Service offers the Carestream In-Vivo Xtreme animal imaging system, which allows up to five mice to be visualized simultaneously using fluorescent and bioluminescent methods. Equipment Carestream In-Vivo Xtreme Multimodal Imaging System Specifications

Back-illuminated 4 MP camera f/1.1 – f/16, 58 mm fixed lens (on movable platform) 400W Xenon Light Source 28 Excitation Filters 410 nm – 760 nm Fluorescent imaging from the visible to near IR Imaging chamber for up to five mice at a time with

integrated anesthesia and temperature control Automated switching between

modalities – the animal never moves Multimodal Animal Rotation System (MARS) Integrated high resolution X-ray imaging (>18lp/mm spot

size <60µm) X-ray illumination optimized for small animal imaging with 5

selectable filter settings for fine tuning Applications

Tumor and Stem Cell Tracking Monitoring of Drug Delivery and Targeting High throughput Pharmacodynamic studies Imaging and validating new Probes and Biomarkers Bionand offers a range of complimentary services including

animal housing facilities, and histology, microscopy and cytometry services

The Magnetic Resonance Service (NMR) is equipped with two High resolution NMR Spectrometers: a 400MHz NMR spectrometer for routine experiments and a higher filed, 600 MHz, spectrometer dedicated to high resolution magic angle spinning (HR-MAS) NMR and in vitro studies that are very demanding in terms of sensitivity. The service also has a TD-NMR analyzer for relaxivity measurements at clinical field (1.44 T).

MAGNETIC RESONANCE SERVICE

Magnetic Resonance Service

HIGH RESOLUTION LIQUID NMR

Liquid NMR is a fundamental tool both in chemistry and biomedicine research. Its applications to chemistry range from structure elucidation of small molecules to studies of ligand-protein interactions. In biomedicine, the use of liquid NMR has expanded from the classical metabolic studies using enriched stable isotope tracers to the more recent NMR-based metabolomics of biofluids (urine, plasma, CSF). Equipment

Bruker Ascend 400 MHz NMR spectrometer

Bruker Avance 600 MHz NMR spectrometer Specifications Bruker Ascend 400 MHz NMR spectrometer

UltraShieldTM Plus 9.4T magnet 5 mm BBFOPLUS probe, optimized for X-nuclei direct

observation. Broad band in a frequency range between 31P and 15N. This probe has 2H “lock” channel and z gradient that allows 2D spectroscopy and hetero correlation 19F/1H and 1H/19F, to be performed. It also includes an automated tuning and matching device (ATM)

Sample Xpress Lite (for 16 samples) Control temperature unit (from 0 ºC to 50 ºC)

Contact Dr. Carmen Muñoz Advanced NMR Specialist Phone +34 952 367 634 Ext. 637 E-mail [email protected]

Specifications Bruker Avance 600 MHz

UltraShieldTM Plus 14.1 T magnet 5 mm BBFOPLUS probe, optimized for X-nuclei direct

observation in a fre-quency range between 31P and 15N. Automated tuning and matching (ATM).

5 mm TXI probe, optimized for higher sensitivity 1H-nuclei observation. The outer coil is optimized for decoupling of 13C and 31P.Automated tuning and matching device (ATM).

TCI CryoProbe Prodigy, uses nitrogen-cooled RF coils and preamplifiers for 1H, 2H and 13C, delivering a 2.5x 1H sensitivity boost. The probe is ATM compatible and has 2H “lock” channel and Z-gradient.

Control temperature unit (from 0 ºC to 50 ºC) Applications

Structure verification and elucidation Molecular mobility: determination

of conformation in solution Determination of enantiomeric excess Kinetic and temperature studies of reaction mixtures Metabolomics of liquid samples (CSF,

plasma, urine and tissue extracts) In vivo Metabolism using enriched stable isotope tracers


HIGH RESOLUTION MAGIC ANGLE SPINNING (HR-MAS) NMR

HR-MAS NMR is based on solid-state NMR, which uses magic-angle spinning at high speeds to average anisotropic interactions, leading to narrower widths. The moderate spin rates used in HR-MAS allows high resolution spectra to be obtained from intact tissue, i.e., without the need of any metabolite extraction procedure. Equipment Bruker Avance 600 MHz NMR spectrometer + HR-MAS Probe Applications

Metabolomics of intact tissue samples (tumor biopsies, …) Structure Characterization and Elucidation

of partially insoluble compounds)



TIME DOMAIN NMR (TD-NMR)


The Minispec MQ-60 provides T1 and T2 measurements with high accuracy and reproducibility, and, most importantly, at clinical field, which is fundamental for translational research. Hence, the Minispec MQ-60 plays a key role in the characterization of new contrast agents for clinical use. Equipment Bruker Minispec (TD-NMR) analyzer Specifications

Permanent 1.4T magnetic field. “Intelligent” 1H probe (“plug and play”)

with 60 MHz operating frequency Fast magnet warm-up and precise digital temperature

regulation for ultimate results stability Optical detector to any kind of glass, quartz and plastic

samples. Specific filters for natural and fluorescent light suppression and energy saving lamps

Applications

Measurement of Relaxation Times (T1, T2, T2*)

The Advanced Fluorescence Service hosts an excellent range of facilities capable of supporting high quality biomedical research. For investigators at Bionand these applications can range from the initial characterization of the fluorescent properties of novel nano-compounds to the evaluation of their medical applications using tissue culture and animal models. For external researchers the range of possible applications goes far beyond nanomedicine to include the characterization of patient tissues, preclinical studies using rodent models, small scale drug screening and studies using model organisms such as Zebrafish or Drosophila Melanogaster. The service is operated in close collaboration with the other advanced facilities of the Nanoimaging Unit, and the other units of Bionand, to ensure that researchers can easily combine fluorescence-based techniques with other complementary approaches such as electron microscopy or nuclear magnetic resonance.

ADVANCED FLUORESCENCE SERVICE

Advanced Fluorescence SERVICE

CONFOCAL MICROSCOPY


Laser-scanning confocal microscopy is a very popular technique that uses a combination of laser illumination and a "pinhole" mask to ensure that only fluorescence from the focal plane reaches the detector. This avoids the characteristic blur typical of conventional fluorescence microscopy and allows images to captured as detailed optical slices and then used to build up rich 3D models. Confocal microscopy is one of the most versatile techniques available in optical microscopy. The Advanced Fluorescence Service offers two extremely well specified Leica SP5 confocal microscopes. Their features include HyD hybrid detectors for the best possible sensitivity, as well as full environmental control for live cells and high-speed resonant scanning. Equipment Leica SP5 HyD Confocal Microscope Specifications

Inverted Leica DMI 6000 fully motorised microscope Confocal detectors: 2 x HyD, 3 x PMT + 1

transmitted light detector Tandem Scanner – HQ conventional scanner and high-speed

Resonant scanner (28 frames/sec @ 512 x 512) AOBS (Acousto-Optical beam splitter) – brighter

and more flexible than con-ventional beam splitters Optical zoom up to 64x (maximum resolution

is dependent on the objective used)

Specifications

Choice of objectives: - Plan Apo Oil immersion: 100x (1.4 NA), 63x (1.4 NA Blue), 40x (1.3 NA) - Plan Apo 20x (0.7 NA) multi-immersion (water, glycerol, oil) 0.26mm WD - Plan Apo 10x (0.4 NA) dry

9 Laser lines in the visible range: - Violet 405nm (50mW), Blue Argon 458nm, 476nm, 488nm, 496nm, 514nm (65mW) Green DPSS 561nm (20mW), Orange HeNe 594nm (2mW), Red HeNe 633nm

Microscope enclosure with temperature, humidity and CO2 control

High precision Super Z motorized stage with multi-point and multi-tile soft-ware control. 3nm resolution galvometric z-axis control

Applications

High-speed live cell imaging: suitable for a wide range of samples including cultured cells and model organisms such as Drosophila Melanogaster

High quality imaging of fixed multi-color immunofluorescence preparations

Small to medium scale screening applications: useful where the highest possible imaging quality is required to detect a particular phenotype

Advanced fluorescence techniques: including FRAP for studying molecular dynamics, photoactivation, and FRET for studying molecular interactions at sub –nanometric distances


MULTIPHOTON MICROSCOPY


Multi-photon (or two-photon) microscopy takes advantages of the near simultaneous absorption of two or more photons which act to excite a fluorescent molecule with the combined energy of both photons. In practice this means that low energy infrared light can be used to see fluorescent molecules that are normally excited by high energy ultraviolet and blue light. Using this method we can visualize fluorescent molecules at much greater depths than conventional microscopy. Infrared light is also much less damaging to live tissues than UV or blue ex-citation, making it ideal for timelapse imaging of model organisms or tissue explants. The Leica SP5 MP HyD combines both confocal and multi-photon modes in a single machine and can easily switch between different imaging methods. It is also fitted with the latest super-sensitive external HyD hybrid detectors dedicated for multi-photon microscopy. Equipment Leica SP5 HyD MP Multiphoton Confocal Microscope Specifications

Inverted Leica DMI 6000 fully motorised microscope Confocal detectors: 2 x HyD, 3 x

PMT + 1 transmitted light detector Tandem Scanner – HQ conventional scanner and high-speed

Resonant scanner (28 frames/sec @ 512 x 512) AOBS (Acousto-Optical beam splitter) – brighter

and more flexible than con-ventional beam splitters Optical zoom up to 64x (maximum resolution

is dependent on the objective used)

Specifications

Choice of objectives: - Plan Apo Oil immersion: 100x (1.4 NA), 63x (1.4 NA Blue), 40x (1.3 NA) - Plan Apo 20x (0.7 NA) multi-immersion (water, glycerol, oil) 0.26mm WD - Plan Apo 10x (0.4 NA) dry - Plan Apo 25x (0.95 NA) water immersion, IR corrected for multiphoton, 2.2mm WD

9 Laser lines in the visible range: - Violet 405nm (50mW), Blue Argon 458nm, 476nm, 488nm,

496nm, 514nm (65mW) Green DPSS 561nm (20mW), Orange HeNe 594nm (2mW), Red HeNe 633nm.

High Performance Mai-Tai titanium-sapphire IR femtosecond pulse laser for multi-photon imaging, tunable from 690-1040nm (maximum power 800nm∼ 2.9W)

Microscope enclosure with temperature, humidity and CO2 control

High precision Super Z motorized stage with multi-point and multi-tile soft-ware control. 3nm resolution galvometric z-axis control

Applications

Deep tissue imaging: imaging thicker tissue sections (>100 microns) and complex models such as Zebrafish where GFP is used as a marker

Long term live imaging: where UV or Blue illumination would be toxic (IR light is much less damaging)

High resolution intravital imaging: Often used in near -surface tissues such as skin, retina and brain

Quantitative photoactivation: multiphoton-excitation only activates mole-cules in the focal plan unlike conventional fluorescence

Fluorescence properties of novel materials: For example nanoparticles often exhibit special absorbance and fluorescence properties when excited by two photon activation


SUPER-RESOLUTION MICROSCOPY


Fluorescence microscopy offers a wide array of tools for visualizing microscopic events. The intrinsic diffraction of light has historically made it difficult to use fluorescence to distinguish structures closer than 200nm apart. Super-resolution microscopy refers to techniques that selectively activate fluorescent molecules to map their position with up to 10 times more accuracy than conventional fluorescent microscopy. With the aim of promoting the use nanoscale imaging in biomedicine, Bionand has established Spain’s first N-STORM super-resolution platforms dedicated to practical applications. The Nikon N-Storm system is capable of localizing molecules with a resolution of up to 20nm.

Equipment Nikon Eclipse Ti N-Storm Specifications

Inverted Nikon Ti Eclipse fully motorised microscope. Andor iXon3 897 EM-CCD camera 100x Plan Apo Oil immersion Objective 1.49 NA 4 Lasers in the visible range:

- Violet 405nm - Argon-Ion 457nm, 477nm, 488nm, 514nm - Yellow Diode 561nm - Red 647nm (300mW)

High precision motorized stage. TIRF optical system (used to improve

contrast with N-STORM samples) Removable cylindrical lens for 3D super-resolution. Nikon PFS (Perfect Focus System) for improved stability NIS-Elements with N-STORM acquisition Offline workstation with additional processing module

Applications

Structure determination of nanometric structures: e.g. nuclear pore receptors where individual subunits can be differentially labeled

High resolution neuron tracing: Super-resolution imaging has been shown to greatly improve the accuracy of neural maps given then many neural projects are below limit of conventional microscopy

Correlative microscopy: the higher resolution of super-resolution fluores-cence makes it easier to associated with structures detected by electron microscopy

Single molecular studies: e.g. synapse neuroreceptor distribution

Nanoscale cytoskeletal and vesicle trafficking studies


FLOW CITOMETRY/ CELL SORTER


Flow cytometry is a highly effective method for analyzing thousands cells or other particles in a matter of minutes. Flow cytometers detect the fluorescence, size and complexity of particles that pass through laser beams as a stream. In this way they can quickly generating statistically valid data by counting very large numbers of particles - something that would be extremely difficult with traditional microscopy approaches. Flow cytometers are very well established in hospitals for routine diagnosis and now widely used for different types of research application, from marine biology to stem cell research. A Cell Sorter is a specialized flow cytometer which is able to physically separate different classes of particles based on their fluorescence or other physical characteristics. Equipment

Beckman Coulter Galios flow cytometer Beckman Coulter MoFlo cell sorter

Specifications

Beckman Coulter Galios flow cytometer Solid State Diode Lasers: Blue 488nm (22mW),

Red 638nm (25mW), Violet 405 (40mW) 10 independent fluorescence channels

with easily interchangeable optical filters Suitable for common fluorescence markers

including: FITC, PE, ECD, PC5, PC5.5, PE-Cy7 125μm spatially separated beam spots Gel coupled 1.2 NA optical collection lens Forward scatter detector with adjustable collection

angle for sub-micron, small and large particles sizes Side scatter detector with independently focused high

performance photodiode and electronic attenuation

Five different signals available from each detector: Integral linear and loga-

rithmic, Peak linear and logarithmic and True Time of Flight linear Time, Ratio User selectable flow rates (Low, Medium and High) 24-tube multi-loader carousel with integrated vortexer Permits analysis of up to 88 tubes an hour at 10,000 events per second

Beckman Coulter MoFlo cell sorter

Validated for 70,000 sorts per second with > 99% purity at all data rates Proven 4-Way Sort for multiple population sorting Independent control of each stream via touch screen control panel High power solid state 488nm and 638nm lasers

Applications

Detection of rare events: the high throughput of Gallios allows for the statically-valid detection of very rare cellular events

Multi-parameter assays: Three lasers and ten channels of Gallios allows the design of complex assays using multiple markers

Cell cycle analysis: detection of DNA content, DNA damage, apoptosis and cell cycle antigens and many others

Marine biology: flow cytometry is increasingly used as a tool to study marine microorganisms using their innate fluorescence properties combined with size and complexity characteristics.

Single cell separation: Mo-Flo is able to automatically sort single cells into individual wells of a 96-well plate with a very high degree of accuracy and is suitable for colony formation, stem cell isolation etc.

Cell purification from tissues: Mo-Flo allows individual cell types from complex tissues to be separated based on size, complexity or markers such as GFP. The yield and purity can be adjusted to suit different applications. Measurement of Relaxation Times (T1, T2, T1)


FLUORESCENCE MICROSCOPY


For many applications conventional widefield fluorescence microscopy is still the best choice. CCD and CMOS-based sensors used for conventional microscopy are often more sensitive that the photomultiplier tubes used in confocal microscopes and flow cytometers. As the camera captures the whole field of view at the same time, it also allows for faster imaging in many cases. Examples where conventional microscopy may be appropriate include the visualization of individual molecules, receptors or small organisms such as bacteria and yeast. Total Internal Reflection Fluorescence (or TIRF) is a powerful technique which combines the sensitivity of conventional fluorescence with selective illumination to improve contrast of features very close to the sample coverslip. TIRF is often used for studies related to membrane dynamics, receptor-ligand interactions and vesicular transport. Users have access to a dedicated conventional fluorescence system equipped for time-lapse microscopy and a laser-based multi-color TIRF system (also used as the base for the N-STORM system). Equipment

Nikon Eclipse Ti basic Fluorescence Microscope Nikon Eclipse Ti TIRF Fluorescence Microscope

Specifications

Nikon Eclipse Ti basic Fluorescence Microscope Inverted Nikon Ti Eclipse fully motorised microscope Hamamatsu Orca R2 monochrome 1.3 MP CCD camera Okolab H101 stage-top incubator with

temperature, humidity and CO2 control Objective lenses:

- Plan Apo Oil immersion: 60x (1.4 NA) - Super Plan Fluor 40x (0.6 NA) Extra Long working distance (2.8-3.6mm) - Plan Apo 10x (0.45 NA)

Applications

Endocytosis and vesicle dynamics: High contrast, speed and sensitivity makes it possible to follow live processes close to the membrane with TIRF

Single molecule studies: The advanced EM-CCD camera allows the fluorescence of individual molecules to be detected even with relatively short exposures

Actin cytoskeleton and focal contact dynamics in live cells: Using fluorescent protein markers these processes can be followed using high time-lapse imaging

Yeast and bacteria studies: It is often difficult to fluorescence from microorgan-isms by confocal microscopy without bleaching, whereas the combination of a 100x 1.49NA objective and a very sensitive camera allows fluorescence to be de-tected without need high laser power or very low exposures.

High speed imaging of calcium dynamics

Nikon Intensilight 130w Mercury Lamp Filter sets for DAPI, GFP, TRITC Nikon PFS (Perfect Focus System) for improved stability

Nikon Eclipse Ti TIRF Fluorescence Microscope

Inverted Nikon Ti Eclipse fully motorised microscope. Motorised TIRF mirror Multipass TIRF dichroic filter (allows

high-speed multicolour TIRF) Andor iXon3 897 EM-CCD camera Microscope enclosure with temperature,

humidity and CO2 control Objective lenses:

- Plan Apo Oil immersion: 100x (1.49 NA) TIRF - Plan Fluor 40x (0.75 NA) - Plan Apo 10x (0.45 NA).

4 Lasers in the visible range: - Violet 405nm - Argon-Ion 457nm, 477nm, 488nm, 514nm - Yellow Diode 561nm - Red 647nm

High precision motorized stage. Nikon PFS (Perfect Focus System) for improved stability

A wide range of other complementary services are available at Bionand including tissue culture and histology services.

Advanced microinjection facilities are also available at Bionand, featuring high performance Eppendorf injection systems combines with inverted Nikon Ti fluorescence microscopes. Contact us for more information.


HIGH CONTENT SCREENING


High content screening (HCS) attempts to bridge the gap between microscopy and cytometry, providing both large quantities of statically useful and phenotypically rich data. High content screening combines automated microscopy with customizable algorithms able to detect and quantify many different types of phenotypic information in both fixed and living cultured cells. The Perkin Elmer Operetta HCS system combines the quality of a spinning disc confocal microscope with the Harmony HCS software, which contains many common algorithms such as nuclei and cells detection, as well as the ability to "learn" new phenotypes. Bionand also offers access to the advanced server-based Columbus HCS software. Equipment Perkin Elmer Operetta HCS confocal Specifications

Spinning disc confocal technology 10x, 20x and 40x long working distance objectives 20x high NA objectives Temperature control and CO2 for cultured cells 400W Xenon based illumination Fully automated image acquisition

from 24,96 and 384 well plates Harmony: integrated acquisition and analysis software Phenologic - machine learning-based

phenotype recognition software Columbus Image Data Storage and Analysis System

Applications

Drug dose response curve calculation: On a single plate multiple concen-trations or combinations of compounds can be easily and quickly tested. The software includes all the analysis and presentation tool required to calculate IC50 values. And display response curves

Small molecule screens based on changes in cell phenotype: Not all phenotypes can be detected easily by cytometry. Operetta allows very complex phenotypes (e.g. organelle distribution or cell organization changes) to be detected automatically and reliably.

Large scale siRNA screens: Automatic detections of up to 384 different siRNAs on a single plate and analysis them automatically makes it possible to perform genome-scale screens for phenotypes of interest

Long term cell culture studies: Operetta is equipped with temperature and CO2 control making it suitable for maintaining live cultured cells in optimal conditions during image acquisition

The Electron Microscopy Service offers access to transmission electron microscopy (TEM), Cryo-TEM and Electron Tomography, high resolution scanning electron microscopy (SEM) and environmental SEM. Special attention has been paid to high-end cryo-inmobilization sample preparation techniques. All the equipment has been configured for best results with biological and nanomedicine applications.

ELECTRON MICROSCOPY SERVICE

Electron Microscopy SERVICE

TRANSMISSION ELECTRON MICROSCOPY (TEM)

Contact Dr. Juan Félix López Téllez Electron Microscopy Specialist Phone +34 952 367 627 E-mail [email protected]

Conventional TEM provides structural information of samples prepared to electron transparency in the sub-nanometer range. Once prepared, many biological samples, including bacteria, viruses, yeasts, cells, and tissue sections can be characterized using TEM. Moreover, TEM can be used to characterize many other types of material such as substrates or nanoparticles making it a highly versatile technique. Equipment FEI Tecnai G2 20 TWIN Transmission Electron Microscope Specifications

Electron source with LaB6 emitter and flexible high tension between 20 and 200 kV

Twin objective lens for high contrast and high tilt applications Side-mounted CCD Olympus Veleta with 2k x 2k MP, high field of

view and fast read-out rates for real time observation Bottom mounted CCD FEI Eagle, 4k x 4k MP, HS scintillator EDS thanks to EDAX DPP-II analyzer with Si (Li) detector Cryo-TEM Electron tomography Sample holders:

- Single tilt - Low background (for EDS applications) double tilt - Fischione 2020 advanced tomography holder - Gatan 626 single tilt liquid nitrogen cryo transfer holder - Gatan 650 single tilt rotation analytical holder


CRYO-TEM


Cryo-TEM is the technique of choice for the visualization of biological suspensions and any other sub-micron particles in solution. Cryo-immobilization is performed through plunge freezing where samples are transferred to the cryo-holder and to the microscope at temperatures close to liquid nitrogen. Visualization with Low Dose mode allows beam damage and re-crystallization risks to be minimized. Equipment Cryo-TEM on FEI Tecnai G2 20 TWIN Transmission Electron Microscope

Gatan Turbo Pumping Station with Cryo-Holder

Gatan Cryo Transfer Workstation

FEI Vitrobot


SCANNING ELECTRON MICROSCOPY (SEM)


SEM comprises a set of techniques related to the interaction of an electron beam as it scans the surface of a sample. Secondary electrons (SE), backscattered electrons (BSE), X rays, Auger electrons and others elements are generated and provides information about surface topography, composition and other properties of the sample. The inclusion of a Field Emission Gun (FEG) source allows resolutions below 2 nm. Equipment FEI Quanta 250 FEG Scanning Electron Microscope Specifications

Everhart-Thornley detector of secondary electrons for high vacuum applications providing topographical information

High contrast backscattered electron detector optimized to work with low volt-ages: vCD detector. Suitable for high-vacuum and low vacuum modes, provides topographical and compositional contrast

Scanning-transmission electron microscopy detector (STEM) for high vacuum applications. Allows to introduce 8 grids in one vacuum cycle and to perform bright field and dark field analysis

Large field detector (LFD) for low vacuum and ESEM applications. Allows working in SE and BSE modes and facilitates the observation of large fields of view

Gaseous secondary electrons detector (GSED) for environmental-SEM (ESEM) applications

Gaseous analytical detector (GAD) for ESEM applications Wet scanning-transmission electron microscopy

detector (Wet-STEM) for ESEM applications


Environmental SEM allows the observation of samples in a vacuum-free atmosphere. Water vapor is introduced into the chamber during the pumping process, allowing to reach chamber pressures as high as 2700 Pa. This pressure combined with a Peltier stage allow us to work at 100% humidity and thus be able to visualize fully hydrated samples and perform dynamic experiments through pressure changes, including hydration - dehydration sequences. Equipment ESEM mode in FEI Quanta 250 FEG Scanning Electron Microscope Specifications

Cooling stage that allows to work In the range between -25 / +55 °C

GSED with a 500 µm aperture, working at pressures as high as 2700 Pa, pro-vides topographical information in low vacuum and ESEM modes

GAD, with a 500 µm PLA cone that extends down 8.5 mm from the unit to allow EDS analysis while BSE detection

LFD can be used simultaneously with GAD and EDX Wet-STEM: in combination with the cooling stage allows

the characterization of wet samples through STEM in ESEM mode

PLA cones for EDS and low kV analysis in ESEM mode


APPLICATIONS OF ELECTRON MICROSCOPY

CELLULAR CHARACTERIZATION Cell structure and ultrastructure visualization and its relationship to function are the most important contributions of EM to cell biology. This includes not only standard visualization of embedded and stained sections but also localization of proteins inside cells through immunolabelling with a resolution around 10 nm. For this high resolution analysis, sample preparation methods that preserves the ultrastructure are essential. High pressure freezing is the cryo-immobilization technique preferred for these kind of characterizations. Moreover, electron tomography is uniquely suited to obtain 3D reconstructions of cells and its internal structures and thus, improve the information provided by standard 2D analysis. LFD can be used simultaneously with GAD and EDX

TISSUE CHARACTERIZATION electron microscopy has a role in the characterization of interactions between cells and with other components inside tissues. Moreover, high resolution surface characterizations of bones, teeth, skin, etc, are typically performed by SEM

OTHER BIOLOGICAL SYSTEMS leaves, flowers, insects, microorganisms are among the structures that can be topographically characterized using SEM. For these systems as for others with high water content, ESEM has become an important tool due to its ability for surfaces visualization even in a 100% humidity environment, reducing sample preparation and permitting dynamic analysis through hydration-dehydration cycles. Dynamics experiments are useful in food industry, water-oil emulsions characterization and pharmaceutical quality control

BIOMATERIALS CHARACTERIZATION electron microscopy plays a double role in biomaterial characterization. On one hand it provides structural and compositional information of the materials engineered to be used inside biological systems, favoring its development. On the other hand, it allows us to visualize their interactions. Functionalized polymeric and metallic nanoparticles for drugs delivery, dental implants, bone plates and cements, artificial tissues are among the biomaterials that can benefit from information provided by EM

MACROMOLECULAR COMPLEXES CHARACTERIZATION structural biology can benefit from electron microscopy to determine 3D structures of macromolecular com-plexes. Negative staining was typically employed in the past but it has been the development of cryo-electron microscopy in combination with Single Particle analysis that has made EM such a very valuable tool for 3D characterization

The construction of the Andalusian Centre for Nanomedicine and Biotechnology BIONAND has been co-financed, with a contribution of 70% of the total cost, by the European Fund of Regional Development (EFRD) together with the Ministry of Economy and Competitiveness in the frame of The Spanish National Plan for Scientific Research, Development and Technological Innovation 2008-2011 (record number. IMBS10-1C-247, quantity. 4,979,567.61 €). The Equipment of the NanoImagen Unit has been co-financed by the European Fund of Regional Development (EFRD) together with the Ministry of Economy and Competitiveness in the frame of The Spanish National Plan for Scientific Research, Development and Technological Innovation 2008-2011 (record number. PCT-420000-2010-003, quantity. 4,305,000 €). The Equipment of the Units of BIONAND has been co-financed by the European Fund of Regional Development (EFRD) together with the Ministry of Economy and Competitiveness in the frame of The Spanish National Plan for Scientific Research, Development and Technological Innovation 2008-2011 (record number. PCT-420000-ACT2, 2011, quantity. 2.424.447 €).

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SMALL ANIMAL MULTIMODAL€¦ · Imaging Service offers the Carestream In-Vivo Xtreme animal imaging system, which allows up to five mice to be visualized simultaneously using fluorescent