For samples with thickness < few microns, such as isolated particles including proteins, macromolecular complexes, decorated liposomes, viruses, bacteria, eukaryotic cells….
Samples are deposited or grown directly onto EM grids and cryo-fixed in their hydrated state by plunge freezing in liquid ethane at liquid nitrogen temperature. As a result, the sample is vitreous (non-crystalline) and embedded in a thin layer of vitreous ice, that ensures the highest structural preservation of the sample.
For imaging in a cryo-TEM, the ice thickness should not exceed ~500 nm.
For thicker samples (> 1 um), three options are available:
- plunge-freezing followed by cryo-FIB-SEM thinning with cryo-lamellae
- HPF followed by cryo-sectioning (CEMOVIS)
- Vitrification by plunge-freezing of Tokuyasu-style Sections (VoS)
Note: Plunge-freezing is compatible with chemical fixation of the sample.
Remodeling of the core leads HIV-1 pre-integration complex in the nucleus of human lymphocytes, J. Virol. 2020 Apr;.