Frozen hydrated bloc-face investigation of tissue for Cryo SEM

Cryo-transmission electron microscopy of frozen-hydrated sections of Escherichia coli and Pseudomonas aeruginosa.

High-pressure freezing of Escherichia coli K-12 and Pseudomonas aeruginosa PAO1 in the presence of cryoprotectants provided consistent vitrification of cells so that frozen-hydrated sections could be cut, providing approximately 2-nm resolution of structure. The size and shape of the bacteria, as well as their surface and cytoplasmic constituents, were nicely preserved and compared well with ot...

Cryo-SEM of Perpendicular Cross Freeze-Fractures Through a High-Pressure-frozen Biofilm

The cryo scanning electron microscopy (cryo-SEM) as well as high-pressure-freezing (HPF) belongs to cutting edge techniques in electron microscopy of hydrated samples [1]. However, their combination is not always easily applicable. Here, we present a way of combining high-pressure-freezing using EM PACT2 (Leica Microsystems) that fixes hydrated samples on a 1.4 mm sapphire discs and high resolu...

Focused ion beam milling of vitreous water: prospects for an alternative to cryo-ultramicrotomy of frozen-hydrated biological samples.

The feasibility of using a focused ion beam (FIB) for the purpose of thinning vitreously frozen biological specimens for transmission electron microscopy (TEM) was explored. A concern was whether heat transfer beyond the direct ion interaction layer might devitrify the ice. To test this possibility, we milled vitreously frozen water on a standard TEM grid with a 30-keV Ga(+) beam, and cryo-tran...

Electron microscopy of frozen hydrated biological specimens.

The use of frozen hydrated specimens for molecular structure determination is limited primarily by radiation damage. The radiation damage effect in frozen hydrated catalase crystals has been measured in terms of the loss of electron diffraction. The results show an improvement for this type or:specimen relative to wet hydrated or to glucose embedded catalase crystals at room temperature. Bright...

Direct SEM Study of Frozen Inner Ear