Instruments and Equipment

Overview

The IBS/ISBG electron microscopy platform is equipped with transmission electron microscopes, cryogenic imaging systems, a cryo-focused ion beam/scanning electron microscope, and dedicated sample-preparation instruments.

Together, these instruments support a broad range of applications, from negative staining and conventional cellular electron microscopy to single-particle cryo-EM, cryo-electron tomography, MicroED, CEMOVIS, cryo-FIB milling, correlative microscopy, and high-pressure freezing.

Instrument selection and sample-preparation workflows are adapted to the specimen, the scientific objective, and the level of structural preservation or resolution required.

Electron Microscopes

Tecnai Spirit T12

Purchased: 2012

Specifications

Accelerating voltage: 120 kV
Electron source: LaB₆
Orius 1000 wide-field camera
Bottom-mounted TVIPS F416 camera, 4K × 4K

Credit: A Delos CEA.

Main Applications

The Tecnai Spirit T12 is primarily used for routine room-temperature electron microscopy, including:

Quality control by negative staining
Morphological analysis of purified macromolecules, complexes, nanoparticles, and viruses
Preliminary single-particle analysis
Conventional cellular electron microscopy
Observation of resin-embedded ultrathin sections
Immunolabelling experiments

Its wide field of view makes it particularly suitable for sample assessment, screening, and conventional ultrastructural analysis.

Tecnai F20

Purchased: 2012

Specifications

Accelerating voltage: 200 kV
Electron source: C-FEG
CETA camera
ASI hybrid detector, 1K × 1K
SerialEM acquisition software
Gatan 626 cryogenic specimen holder
Gatan 614 cryo-tomography holder

Credit: A Delos CEA.

Main Applications

The Tecnai F20 is a versatile cryo-equipped transmission electron microscope used for both room-temperature and cryogenic imaging.

Applications include:

Negative staining
Cellular electron microscopy
Cryogenic grid screening
Single-particle cryo-EM
Cryo-electron tomography
Observation of CEMOVIS sections
Tomography of cellular and molecular specimens
Microcrystal electron diffraction, or MicroED

The side-entry cryogenic holders allow vitrified specimens to be observed and tilt series to be acquired for tomographic reconstruction.

Thermo Scientific Glacios Cryo-TEM

Purchased: 2019

Specifications

Accelerating voltage: 200 kV
Electron source: X-FEG
CETA camera
Gatan K2 Summit direct electron detector
Falcon 4i direct electron detector
Automated cryogenic specimen loading

Available acquisition and imaging software includes:

EPU 3 with multigrid acquisition
Tomo 5
MAPS
Latitude S
SerialEM

Main Applications

The Glacios is the platform’s principal instrument for cryogenic grid screening, sample optimization, and 200 kV cryo-EM data collection.

It is used for:

Screening grids before high-resolution data collection on a Titan Krios
Assessment of ice thickness and grid quality
Evaluation of particle concentration and distribution
Identification of aggregation or preferential particle orientation
Automated single-particle data collection
Molecular and cellular cryo-electron tomography
Collection of preliminary datasets to demonstrate sample suitability
High-resolution single-particle structure determination

Depending on the specimen and grid quality, the microscope can support near-atomic-resolution single-particle reconstructions. The platform has achieved reconstructions at resolutions down to approximately 2.2 Å.

The Glacios is also an important intermediate step between initial vitrification and advanced data collection on a 300 kV Titan Krios. Thermo Fisher Scientific similarly positions the Glacios as a 200 kV cryo-TEM for high-performance single-particle analysis, tomography, and pre-screening within integrated Glacios–Krios workflows.

Cryo-FIB/SEM

Thermo Scientific Aquilos 2 Cryo-FIB

Purchased: 2025

Specifications

Dual-beam cryogenic FIB/SEM system
Electron-column accelerating voltage up to 30 kV
X-FEG electron source
Gallium focused-ion-beam column
Cryogenic specimen stage
Integrated fluorescence microscope with four fluorescence wavelengths
Capability for cryogenic lift-out experiments
Automated or semi-automated lamella-milling workflows

Main Applications

The Aquilos 2 is dedicated to the preparation of thin, electron-transparent cryogenic lamellae from vitrified biological samples.

Applications include:

Cryo-FIB milling of cells grown on electron microscopy grids
Preparation of lamellae from bacteria, organelles, tissues, and other relatively thick specimens
Targeted milling guided by fluorescence microscopy
Preparation of samples for cellular cryo-electron tomography
Cryogenic lift-out from bulk-frozen specimens
Preparation of lamellae for MicroED
Correlative cryogenic fluorescence, electron, and ion-beam imaging

In a typical workflow, a vitrified cell or biological specimen is first imaged by scanning electron microscopy. Fluorescence information may then be used to identify a region of interest. Material surrounding this region is progressively removed with the focused gallium ion beam until a thin lamella suitable for cryo-TEM imaging and tomography is obtained.

The Aquilos 2 is specifically designed for the preparation of electron-transparent lamellae for high-resolution cellular cryo-electron tomography. Its integrated fluorescence capabilities facilitate correlation and targeting without requiring repeated transfers between independent imaging systems.

Cryo-EM Grid Preparation

Glow-Discharge Devices

The platform is equipped with EMS and GloQube glow-discharge systems for preparing electron microscopy grids.

Glow discharge exposes the grid surface to a low-pressure plasma. This treatment cleans the support film and modifies its surface properties. Carbon-coated grids, which are naturally relatively hydrophobic, can thereby be made more hydrophilic so that aqueous biological samples spread more evenly over the grid.

Glow discharge can improve:

Grid wettability
Distribution of aqueous samples
Adsorption of proteins or particles
Reproducibility of negative-staining preparation
Reproducibility of cryo-EM grid preparation

EMS Glow-Discharge Device

The EMS system is used for routine air glow discharge of carbon-coated and holey-carbon grids. It provides a simple treatment for cleaning and hydrophilizing grid surfaces before negative staining or vitrification.

GloQube Glow-Discharge System

The GloQube provides controlled and reproducible glow-discharge treatment using programmable protocols. Depending on its configuration, the system can support in-air or chemical-vapour treatments and positive or negative discharge modes.

These options can be useful when different surface charges or hydrophilic properties are required to promote sample adsorption, influence particle distribution, or reduce preferential orientation.

Glow-discharge conditions must be adapted to the support film and sample. Excessive treatment may damage particularly delicate films, while some specialized supports should not be glow discharged.

EMS and Quorum describe these devices as systems for modifying the surfaces of TEM grids to improve the imaging of macromolecular samples. The GloQube platform provides automated, recipe-based control for reproducible surface treatment.

Vitrobot Mark IV

Available systems: Two standard instruments

The Thermo Scientific Vitrobot Mark IV is a semi-automated plunge-freezing system used to vitrify liquid biological samples on electron microscopy grids.

A small volume of sample is deposited onto a grid inside an environmental chamber. Excess liquid is removed by programmable blotting, after which the grid is rapidly plunged into liquid ethane or another appropriate secondary cryogen.

The instrument allows important preparation parameters to be controlled, including:

Chamber temperature
Relative humidity
Blotting time
Blotting force
Number of blotting cycles
Waiting time before blotting
Waiting time before plunging
Grid position relative to the blotting pads

Main Applications

Vitrification of purified proteins and macromolecular complexes
Preparation of grids for single-particle cryo-EM
Preparation of molecular cryo-ET specimens
Vitrification of viruses and virus-like particles
Vitrification of liposomes, vesicles, and nanoparticles
Plunge freezing of small cells or bacteria grown on grids
Optimization of ice thickness and particle distribution

The controlled temperature and humidity chamber helps reduce variability between grids and makes it possible to optimize preparation conditions systematically. Thermo Fisher Scientific describes the Vitrobot Mark IV as a semi-automated system that maintains controlled temperature, humidity, blotting, and freezing conditions for reproducible cryo-EM specimen vitrification.

Vitrobot Mark IV in an Anaerobic Glove Box

A dedicated Vitrobot Mark IV is installed inside an anaerobic glove box.

This configuration enables vitrification while limiting exposure to atmospheric oxygen and is intended for oxygen-sensitive or strictly anaerobic specimens.

Potential applications include:

Oxygen-sensitive enzymes
Redox-sensitive macromolecular complexes
Anaerobic microorganisms
Metalloproteins containing oxygen-sensitive cofactors
Samples whose structure or activity changes rapidly following oxygen exposure

Sample preparation, transfer to the grid, blotting, and plunge freezing can therefore be performed in a controlled low-oxygen environment.

The compatibility of the buffer, sample container, cryogens, and transport procedure with the glove-box workflow should be discussed with platform staff before the experiment.

Cellular Electron Microscopy Sample Preparation

Leica EM UC7–FC7 Cryo-Ultramicrotome

The Leica EM UC7 ultramicrotome is combined with an EM FC7 low-temperature cryo-sectioning chamber.

The UC7 can be used to trim specimens and cut ultrathin sections at room temperature. Addition of the FC7 chamber allows sectioning to be performed at controlled sub-zero temperatures.

Main Applications

Ultrathin sectioning of resin-embedded samples
Semi-thin sectioning for light-microscopy orientation
Tokuyasu cryo-sectioning
Cryo-ultramicrotomy
CEMOVIS preparation
Sectioning of polymers or temperature-sensitive materials
Preparation of sections for immunolabelling
Surface preparation and trimming before imaging

For Tokuyasu applications, chemically fixed and cryoprotected specimens are sectioned at low temperature before immunolabelling. For CEMOVIS, vitrified biological material is cut directly under cryogenic conditions, without conventional resin embedding.

CEMOVIS is technically demanding and requires careful control of sample thickness, vitrification, cutting temperature, knife condition, compression, and section transfer.

Leica describes the FC7 as a low-temperature chamber that converts the UC7 ultramicrotome into a cryo-sectioning system suitable for ultrathin cryo-sections and the preparation of cryogenic material.

Leica EM AFS2 and EM AFS1 Freeze-Substitution Systems

The platform is equipped with Leica EM AFS2 and EM AFS1 systems for freeze substitution and low-temperature sample processing.

Freeze substitution is generally performed after high-pressure freezing or another cryofixation method. While the specimen remains at very low temperature, vitrified water is gradually replaced with an organic solvent, commonly acetone or methanol. Fixatives, stains, or contrasting agents may be incorporated into the substitution medium according to the protocol.

The temperature is then progressively increased before resin infiltration, embedding, and polymerization.

Main Applications

Processing of high-pressure-frozen cells and tissues
Freeze substitution of bacteria, pellets, organoids, and small organisms
Low-temperature chemical fixation
Low-temperature staining and contrasting
Resin infiltration and embedding
Progressive lowering of temperature protocols
Preparation for conventional TEM
Preparation for electron tomography
Preparation of samples for correlative light and electron microscopy

Freeze substitution combines the structural preservation obtained through rapid cryofixation with the practical advantages of resin embedding and room-temperature ultramicrotomy.

The AFS2 allows programmed control of temperature and processing time and supports freeze substitution, progressive lowering of temperature, low-temperature embedding, and resin polymerization. The AFS1 provides additional capacity for established freeze-substitution protocols.

Leica EM GP2 Automatic Plunge Freezer

The Leica EM GP2 is an automatic plunge-freezing system used to vitrify liquid or very thin biological specimens deposited on electron microscopy grids.

The sample is maintained in a protected environmental chamber with controlled temperature and humidity. Excess liquid is removed automatically before the grid is rapidly plunged into liquid ethane.

Main Applications

Preparation of grids for single-particle cryo-EM
Preparation of molecular cryo-ET specimens
Vitrification of viruses and macromolecular complexes
Plunge freezing of bacteria and small cells
Preparation of thin cellular specimens for cryo-electron microscopy
Optimization of ice thickness and sample distribution

The instrument complements the Vitrobot systems and provides an alternative approach to blotting and vitrification. Selection of the most appropriate plunge freezer depends on the grid type, sample behavior, required blotting geometry, and project workflow.

The EM GP2 provides controlled temperature and humidity conditions and automated blotting before plunge freezing into liquid ethane. Leica specifies an environmental chamber adjustable between approximately 4 °C and 60 °C and up to 99% relative humidity.

Leica EM ICE High-Pressure Freezer

The Leica EM ICE is used for rapid cryofixation of biological samples by high-pressure freezing.

During the process, the specimen is exposed almost simultaneously to high pressure and rapid cooling with liquid nitrogen. High pressure suppresses ice-crystal formation and allows thicker specimens to be vitrified more effectively than is generally possible by conventional plunge freezing.

Main Applications

Cryofixation of tissues
Cryofixation of cell pellets
Preservation of organoids and small organisms
Freezing of concentrated cellular or microbial specimens
Preservation of organelles and membrane systems
Time-resolved freezing following light or electrical stimulation
Preparation for freeze substitution and resin embedding
Preparation for CEMOVIS
Preparation of bulk specimens for cryo-FIB lift-out
Correlative light and electron microscopy workflows

High-pressure freezing is particularly valuable when preservation of membranes, organelles, cytoskeletal elements, and other delicate cellular structures is required.

After freezing, specimens may be processed by freeze substitution using the AFS1 or AFS2, embedded in resin and sectioned for conventional TEM. Alternatively, they may be maintained in the vitrified state for cryogenic sectioning, cryo-FIB preparation, or cryo-electron microscopy.

The EM ICE uses a liquid-nitrogen-based, alcohol-free freezing principle and is designed to preserve biological ultrastructure in a close-to-native state. Leica recommends combining high-pressure freezing with downstream workflows such as freeze substitution, cellular electron microscopy, CLEM, and cryogenic analysis.

Integrated Instrument Workflows

The instruments can be combined into several complementary workflows.

Single-Particle Cryo-EM

Sample quality control by negative staining on the Tecnai Spirit T12
Glow discharge of the cryo-EM grid
Vitrification using the Vitrobot Mark IV or Leica EM GP2
Grid screening and optimization on the Glacios
Data collection on the Glacios or a 300 kV Titan Krios

Molecular Cryo-Electron Tomography

Glow discharge and grid preparation
Vitrification using the Vitrobot or EM GP2
Screening on the Tecnai F20 or Glacios
Tilt-series acquisition on the Tecnai F20, Glacios, or Titan Krios
Tomogram reconstruction and, when appropriate, subtomogram averaging

Conventional Cellular Electron Microscopy

Chemical fixation or high-pressure freezing
Freeze substitution using the AFS1 or AFS2, when applicable
Resin embedding
Ultrathin sectioning using the UC7
Imaging on the Tecnai Spirit T12 or Tecnai F20

Cellular Cryo-Electron Tomography

Plunge freezing using the Vitrobot or EM GP2, or bulk vitrification using the EM ICE
Fluorescence localization when required
Lamella preparation using the Aquilos 2 cryo-FIB/SEM
Cryogenic TEM screening
Tilt-series acquisition and tomographic reconstruction

Instrument Access

Instrument access depends on the proposed experiment, sample readiness, biosafety requirements, staff availability, and the level of user training.

Users are encouraged to contact the platform at an early stage so that the most appropriate preparation and imaging workflow can be defined.

Some instruments are operated exclusively or primarily by platform staff. Independent use may require prior training and authorization.

Location

The instruments are located at the Institut de Biologie Structurale in Grenoble, within the Methods and Electron Microscopy Group.

Most electron microscopy and sample-preparation facilities are situated on the ground and first floors of the IBS building.