The construction and operation of a simple inexpensive slam freezing device for electron microscopy

Abstract

SUMMARY A simple and inexpensive slam freezing device has been constructed that allows consistently good rapid freezing of uncryoprotected tissue. The device employs a dry liquid‐nitrogen‐cooled copper surface and a slamming plunger designed to provide rapid and uniform tissue to metal contact.

Immunocytochemical strategies for electron microscopy: choice or compromise

“…This is ample depth for samples of small isolated cells or bacteria but it can be a major limitation for dissected organs. Impact freezing devices are available commercially and there are descriptions of simple home‐made devices in the literature (Heath, 1984; Allison et al ., 1987). The speed of delivery onto the cooling surface or ‘bounce’ may cause tissue damage (Sitte et al ., 1987) and there may well be mechanical distortion of tissue architecture even after optimizing tissue contact with the cooling surface (Bennet, 1998).…”

“…the literature (Heath, 1984;Allison et al, 1987). The speed of delivery onto the cooling surface or`bounce' may cause tissue damage (Sitte et al, 1987) and there may well be mechanical distortion of tissue architecture even after optimizing tissue contact with the cooling surface (Bennet, 1998).…”

Electron microscopy of high pressure frozen samples: bridging the gap between cellular ultrastructure and atomic resolution

“…Ice forming in the cytoplasm induces phase segregation between water and solutes (organic matter and salts), with the eVect that cellular components are concentrated and precipitate between ice crystal ramiWcations. In electron micrographs, so-called segregation patterns become visible (Allison et al 1987;Dubochet 2007;Escaig 1982). In the worst case, growing ice crystals poke holes into cellular membranes and destroy organelles (Meryman 2007).…”

An improved cryofixation method: cryoquenching of small tissue blocks during microwave irradiation

“…In the absence of cryoprotectants, extremely fast freezing rates of > lo4 K/s can minimize ice-crystal damage (Plattner & Bachmann, 1982;Sitte et al, 1986). A number of methods have been developed to increase the freezing rates: spray or immersion freezing in which particulate samples are sprayed or rapidly immersed into liquid cryogen (Bachmann & Schmitt, 1971;Rebhun, 1972;Glover & Garvitch, 1974;Costello & Corless, 1978;Echlin, 1978;Handley et al, 1981;Plattner et al, 1982;Sitte et al, 1986), jet freezing in which liquid propane or cryogen is sprayed onto the sample (Moor et Terracio Escaig, 1982;Bald, 1983Bald, , 1985Sitte et al, 1986;Allison et al, 1987). The high-pressure method greatly improves heat transfer by preventing boiling of the cryogen at the specimen surface, thus depressing the freezing point of water and reducing ice-crystal growth (Bald, 1984;Muller & Moor, 1984) and, so far, seems to be the best for prevention of ice-crystal formation (Muller & Moor, 1984;Kaeser et al, 1989).…”

“…This, together with additional but conventional technical improvements of use, i.e. the light-mass sample holders (Escaig, 1982), and prevention of bouncing (Allison et al, 1987) by the use of a recently developed fl-gel shock absorber, has enabled us to obtain good ultrastructural preservation throughout the specimen up to a depth of 15 pm with excellent accuracy and stability for repetition of the experiments. The improved method, the freezing apparatus and its application to squid retina, rat liver and rat heart muscle are reported in the present paper.…”