Available Services

This service is designed to cover imaging on large volumes and includes sample preparation by either high-pressure freezing or chemical fixation, freeze substitution, resin embedding, section preparation by ultramicrotomy and S\TEM imaging at 300kV. The User can provide specimen already fixed, stained, embedded as well as already sectioned and made ready for EM acquisition. A maximum of 1 specimen can be processed per unit of service, and 4 samples maximum (i.e., replicates) can be prepared per specimen. High-pressure freezing will be performed on a Leica EM ICE. Freeze substitution will be performed on a Leica AFS2, and a User might specify a substitution protocol of preference, which will be utilized if compatible with Facility practices. Similarly, for chemical fixation, the User is allowed to specify their protocol of preference, which will be adopted if compatible with Facility practices. For resin-embedding, the User could provide a resin of preference, or alternatively, the currently available resin be used. Sections of resin-embedded sample will be prepared on Leica UC7. A maximum of 1 resin-embedded sample can be processed for unit of service, to be sectioned in ribbons, and to be applied to a maximum of 4 TEM support grids. User might specify thickness of final sections, and these recommendations will be followed if compatible with the Facility practices. User might provide TEM grid support of preference, otherwise the currently available grid support will be used. Imaging will be performed an a Thermo Scientific Spectra 300 kV either in TEM, equipped with CETA 2 camera, or STEM mode in BF\DF\HAADF. Imaging conditions, if requested by the Usermust be compatible with Facility practices. Microscope access is granted for a maximum of 5 days per unit of service, including all steps from sample loading to alignments and collection, and according to Facility Staff availability. All steps of this service will only be performed by Facility Staff. To maximise efficient use and access to high-end TEM instruments, priority will be given to applicants specifically requesting parts of this service (e.g. only sectioning, or only S\TEM imaging of already prepared sections).

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We offer sample preparation by negative stain EM followed by TEM imaging at 120 kV. A maximum of 1 specimen and 8 grids can be prepared and processed per unit of service. Imaging will be provided for a maximum of 8 continuous hours per unit of service. An optional “polishing” size-exclusion chromatography (SEC) step can be performed on thawed material by the Biophysics Unit, as and if specified in the request for this service. 400 mesh copper grids with amorphous carbon film layer will be used as support. Glow discharging will be performed by a Pelco EasyGlow device. Staining will be performed with a 2% (w\v) uranyl acetate aqueous solution. Imaging in TEM mode at 120 kV will be performed on a Thermo Scientific Talos L120C equipped with CETA 16M camera. Imaging conditions requested by the User, if provided, must be compatible with Facility practices. This service will only be performed by Facility Staff.

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This service provides high-resolution cryo-TEM data collection of vitrified specimens. According to instrument availability and experimental needs, data collection will be carried out either on 200kV or 300kV microscope systems. A maximum of 1 specimen can be imaged per unit of service. To ensure efficient usage of high-end microscope time, a maximum of 4 grids can be loaded per unit of service. The User can provide cryo-TEM grids either unmounted or mounted on a Thermo Scientific cartridge. In case of an unmounted grid, clipping of the specimen in Thermo Scientific cartridges will be performed by Facility Staff. In case of User-provided already-clipped grids, these will be inspected by Facility Staff prior to acceptance. Imaging at 200 kV will be performed on a Thermo Scientific Glacios while imaging at 300 kV will be performed on a Thermo Scientific Titan Krios G4, both equipped with a Falcon 4i direct electron detector and a Selectris X energy filter. Imaging conditions (i.e., dose, pixel size, magnification, etc.), if requested by the User, must be compatible with the Facility best practices. Microscope access is granted for a maximum of 24 continuous hours per unit of service, including all steps from clipping to loading and TEM alignments and according to Facility Staff availability. For single-particle acquisition, User might opt for beam-image shift assisted data collection (~ 450 – 600 movies\hour) or stage movement for each hole (~ 100 – 250 movies\hour). This service will only be performed by Facility Staff.

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This service provides cryo-Fluorescence Microscopy imaging including sample preparation by plunge-freezing and grids clipping. A maximum of 1 specimen can be processed per unit of service, maximum 4 samples (i.e. replicates) can be prepared per specimen. Glow discharging will be performed by either a Pelco EasyGlow, a Quorum GloQube, or Solarus II plasma cleaner device. Plunge-freezing will be performed on a Thermo Scientific Vitrobot Mk IV or a Leica EM GP2. Applicants may provide TEM grid supports of preference, otherwise the Facility Staff will use the available TEM grid supports. Widefield imaging will be performed on a Leica Thunder cryo-CLEM system. Confocal imaging will be performed on a Leica Stellaris 5 cryo-CLEM system equipped with white light laser. Both microscopes are equipped with a 50x \ 0.9 NA lens. This service is provided for a maximum of 8 continuous hours per unit of service, including plunging, clipping, and imaging. In case of User-provided already-clipped grids (in Thermo Scientific cartridges), these will be inspected by Facility Staff prior to acceptance. This service will only be performed by Facility Staff.

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This service provides sample preparation by plunge-freezing, grid clipping for autoloader system and cryo-TEM imaging at 200 kV. A maximum of 1 specimen and 8 grids can be prepared and processed per unit of service. An optional “polishing” SEC step can be performed on thawed material by the Biophysics Unit, as and if specified in the request for this service. Glow discharging will be performed by either a Pelco EasyGlow, a Quorum GloQube, or Solarus II plasma cleaner device. Plunge-freezing will be performed on a Thermo Scientific Vitrobot Mk IV or a Leica EM GP2. Applicants may provide TEM grid supports of preference, otherwise Facility Staff will use holey TEM grid supports available. Imaging will be provided for a maximum of 8 continuous hours per unit of service. Cryo-TEM imaging will be performed only in EF-TEM mode at 200 kV on a Thermo Scientific Glacios equipped with a Falcon 4i direct electron detector and Selectris X energy filter. Imaging conditions requested by the User, if provided, must be compatible with Facility practices. In case of grids showing optimal particle distribution and ice quality, this service can be extended for overnight data collection, if compatible with Facility Staff working hours. The User can also provide already-made cryo-TEM grids either unmounted or mounted on a Thermo Scientific cartridge. In case of an unmounted grid, clipping of the specimen in Thermo Scientific cartridges will be performed by Facility Staff. In case of User-provided already-clipped grids, these will be inspected by Facility Staff prior to acceptance. This service will only be performed by Facility Staff.

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