Jet freezing of cells and tissues with and without cryoprotectants

Abstract

For the past several years cryofixation/freeze-substitution techniques have become valuable alternatives to chemical fixation of biological specimens. The superiority of cryofixation in preserving labile cell structures has been documented in several studies. Commercially available jet freezers and the BAL-TEC HPM010 high pressure freezer have extended high quality cryofixation from monolayer cells to cells relatively deep inside tissues. High pressure freezing can theoretically freeze biological materials of 0.5 mm thickness without the use of cryoprotectants and propane jet freezing is reported to freeze biological samples up to 40 μm in thickness without cryoprotection. Although high pressure freezing is the obvious method of choice for freezing large biological samples, its high cost combined with its apparent inability to consistently preserve microfilaments in some plant cells has prompted us to explore the capability of jet freezing to yield well frozen samples with and without cryoprotectants.We used the commercially available jet freezer JFD 030 (BAL-TEC) to obtain our results. Tightly pelleted cells sandwiched between 0.1 mm thick copper specimen carriers normally froze well without any cryoprotectants, after propane jet freezing (Figs. 1-2).