Hydra Bio Plasma-FIB Applications

Hydra Bio Plasma-FIB applications for volume imaging

Explore across length scale from tissues to proteins with the Thermo Scientific Hydra Bio Plasma-FIB. Investigate subcellular structures within the contextual environment by volume imaging. Study resin-embedded specimens at room temperature and unstained biological samples in the near-native state at cryogenic temperatures. Bridge the gap in terms of specimen volume and resolution between light microscopy and cellular cryo-electron tomography. 

Hydra Bio Plasma-FIB applications in cryogenic volume electron microscopy

Hydra Bio Plasma-FIB applications for volume electron microscopy at room temperature

3D reconstruction of mouse neuron

3D reconstruction of mouse neuron prepared in epoxy resin and imaged with the Hydra Bio Plasma-FIB. Segmentation shows synaptic vesicles and mitochondria (red). 

Mouse hippocampus visualized with the Spin Mill Bio Method

Spin Mill Bio Method allows milling of large areas to reveal high-resolution sample details which can be further analyzed to focus in on regions of interest. Mouse hippocampal organotypic slice in EPON resin. Courtesy of S. Watanabe, John Hopkins University and J. Wang, Thermo Fisher Scientific.

Cross section of a tobacco leaf

Cross section of a tobacco leaf, stained and embedded in Quetol resin. The large field of view is made possible with the Spin Mill Bio Method. Imaging with oxygen rather than gallium reduces charging and avoids curtaining artifacts that would otherwise be experienced with Quetol resin. Sample courtesy of Kirk Czymmek, Donald Danforth Plant Science Center, USA.

Cryo-tomography with the Hydra Bio Plasma-FIB

Cryo-electron tomography (cryo-ET) is a technique that allows the study of the 3D structure of cells and tissues at near-native conditions. Sample preparation is a critical step in cryo-ET, which can greatly affect the quality and resolution of the results. Cryo-ET has been applied to many different sample types, from single molecules to protein complexes, viruses, bacteria, cells, to tissue cells and large tissue samples. Samples are either vitrified through plunge-freezing or high pressure freezing (HPF). After the vitrification step, depending on the size of the sample, a thinning step is included in the sample preparation using (plasma) focused ion beam milling to produce cryo-lamellae that are thin enough to be penetrated by electron beams. The Hydra Bio Plasma-FIB supports automated preparation of lamellae with a thickness of 150 nm or less for the cryo-electron tomography workflow, while avoiding gallium-ion implantation. The guided workflow allows the selection of multiple points of interest and then automatically prepares several lamellae autonomously in unattended runs. The Hydra Bio Plasma-FIB prepares lamellae from within large (HPF) volumes with plasma FIB, CLEM localization through integrated light and electron imaging (iFLM Correlative System), and cryo-lift out with Thermo Scientific Easylift Cryo.

Correlative light and electron microscopy (CLEM)

The Thermo Scientific Integrated Fluorescence Light Microscope (iFLM) Correlative System is an integrated wide-field light microscope for correlative imaging inside the Hydra Bio Plasma-FIB, enabling fluorescent imaging and ion milling within a single microscope at room temperature or under cryo conditions.

AI-assisted serial section electron microscopy with Auto Slice & View Software

Combined with Thermo Scientific Auto Slice & View Software, the Hydra Bio Plasma-FIB offers high throughput and fully automated 3D data collection, even at cryogenic temperatures. Benefitting from real-time AI analysis, dynamic auto functions effectively mitigate the impact of sample variability, which is an essential advantage in unattended, large-volume acquisition of cryo-prepared specimens. 

Lamella preparation with AutoTEM Cryo Software

Thermo Scientific AutoTEM Cryo Software offers the highest level of automation and an intuitive user interface with hints and instructive graphics. This simplifies and shortens the lamella preparation time for experts and enables beginners to routinely obtain the highest quality results.  Create unattended batch milling even for overnight runs with configurable steps from milling to polishing and drift correction.

Array tomography with the Hydra Bio Plasma-FIB

Array tomography can provide rapid localization of cells and their interaction partners in tissues. With the largest imaging field, array tomography is ideal for tissue histology. Ultrastructure analysis of whole animals or whole organs is not needed, so array tomography provides easy navigation for samples that can be stored and re-imaged. Like any Thermo Scientific SEM, the volume EM capability of the Hydra Bio Plasma-FIB can be extended with the addition of automated Maps for Array Tomography Software.

Maps Software for Array Tomography

Thermo Scientific Maps Software is an imaging and correlative workflow software suite compatible with Hydra Bio Plasma-FIB.

Protocol: Targeted 3D EM and Correlative Microscopy Method using SEM Array Tomography

Optimized scanning electron microscopy workflow for volume array tomography for asymmetric samples and model organisms.

Header image courtesy of Tessa Burch-Smith, Kirk Czymmek and Lolita Rotkina, Donald Danforth Plant Science Center.

For Research Use Only. Not for use in diagnostic procedures.