A SIMPLE PNEUMATIC DEVICE FOR PLUNGE-FREEZING CELLS GROWN ON ELECTRON-MICROSCOPY GRIDS

A detailed design for a simple and inexpensive variablespeed (1.05.8 m s1) pneumatic plungefreezing device is presented. Cultured cells, grown on Formvarcoated 75mesh gold finder grids, are pneumatically driven into a stirring mixture of propane/isopentane (3:1) cooled by liquid nitrogen (LN2). Premature freezing of the sample in the cryogenic vapors above the cryogen is prevented by plunging through an entry tube into an insulating box, to which a partial vacuum is applied. The cryogenic vapors are drafted into the box at the level of the liquid cryogen by the vacuum, thereby preventing a layer of cold gas from collecting above the cryogen. To prevent the sample from thawing during transfer from the cryogen to the substitution medium, the box top is removed and compressed air is forced through a corrugated tube running the length of the box. The resulting boiling LN2 creates an atmosphere below120C in which the transfer can be accomplished. Copyright1990 WileyLiss, Inc.