Freeze‐fracturing: a review of methods and results

SUMMARY Freeze‐fracturing may be accomplished either under vacuum, or at atmospheric pressure. The devices available for freeze‐cleaving are discussed in relation to the vacuum and temperature conditions prevailing during cleaving, etching and replication. It is concluded that most specimens can be satisfactorily cleaved (even at 4 K), and processed using simple cleavage devices and systems that provide ample cold‐trapping protection for the specimen. Only for special purposes are microtome assemblies or ultra‐high vacuum units essential. The fracturing process may produce artefacts by plastic deformation. Such artefacts have been noted in both non‐biological and biological polymers cleaved at temperatures as low as 4 K. Contamination of the frozen surface need not be a problem provided that the specimen is transferred under ‘safe’ conditions, and is protected by well‐designed cold traps. Freeze‐cleavage and freeze‐sectioning are compared. It is considered that there will be a temperature range for most heterogeneous specimens within which both cleavage and fracturing may occur, depending upon the nature of the molecules in the cleavage/sectioning plane. Local heating during freeze‐sectioning will produce deformation artifacts as in freeze‐cleavage, and may also lead to more general surface ‘flow’. The relationships between sectioning and fracturing biological specimens at low temperature require further clarification.