PFIB | Hydra Bio Plasma FIB Applications | Thermo Fisher Scientific

Hydra Bio Plasma-FIB uses and applications

Explore tissues to proteins with the Thermo Scientific Hydra Bio Plasma-FIB. Perform volume imaging at room temperature or in native state with excellent contrast on frozen cells and tissues. Biological samples are fixed in near-native conditions and are stain-free. Bridge the gap between light microscopy and cellular cryo-electron tomography, both in terms of volumes that can be interrogated as well as attainable resolution.

Cryogenic volume electron microscopy with the Hydra Bio Plasma-FIB

Volume electron microscopy with the Hydra Bio Plasma-FIB

3D volume electron microscopy of mouse neuron containing synaptic vesicles and mitochondria (colored) HPF in epoxy resin.

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 culture HPF Freeze Resin EPON. Courtesy of S. Watanabe, John Hopkins University and J. Wang, Thermo Fisher Scientific.

Courtesy of Dr. Kirk J. Czymmek, Principal Investigator, Director Advanced Bioimaging Laboratory, 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 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 PFIB 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 Florescent Light Microscope (iFLM) Correlative System is an integrated wide-field light microscope for correlative imaging inside the Hydra Bio PFIB, enabling fluorescent imaging and ion milling within a single microscope at room temperature or under cryo conditions.

Serial section electron microscopy with Auto Slice & View Software

Combined with optional Thermo Scientific Auto Slice & View Software, the Hydra Bio PFIB offers high throughput and fully automated 3D data collection, even at cryogenic temperatures. This automates collection of contextual volume imaging.

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 Hydra Bio FIB can be turned into a volume EM microscope with the addition of automated Maps for Array Tomography Software.