Cryo-preparation is a reliable technique for the structural investigation of food products in low temperature scanning electron microscopy (SEM). Artifacts, such as, the segregation of water/non-water ingredients, occur during the freezing process by the crystallization of ice; they can be helpful for correct interpretation of visualized details, e.g., the detection of water containing compartments. The size of the segregation structures depends on water concentration and specimen thickness. The condensation of water vapor (ice contamination) is influenced by the specimen temperature and the partial pressure of the water inside the vacuum system. Furthermore, the evaporation (sublimation, etching) of specimen water can be regulated by monitoring the specimen temperature. Sublimation under SEM observation, i.e., "in situ etching" at low acceleration voltage, allows the progress of etching to be observed continuously, prior to the coating of the specimen inside a dedicated cryo-preparation system attached to the SEM. Coating of specimens provides superior structural resolution compared with the observation of uncoated samples. A coating layer of platinum ( ~ 1-2 nm thick), deposited on a cold substrate by planar magnetron sputtering, is almost homogenous and has a density close to that of the solid metal. Its use allows bulk biological specimens to be observed in low temperature SEM with a structural resolution up to the visualization of transmembrane proteins.
Müller, T.; Walther, P.; Scheidegger, C.; Reichelt, R.; Müller, S.; and Guggenheim, R.
"Cryo-Preparation and Planar Magnetron Sputtering for Low Temperature Scanning Electron Microscopy,"
Scanning Microscopy: Vol. 4
, Article 6.
Available at: https://digitalcommons.usu.edu/microscopy/vol4/iss4/6