Freeze substitution of high-pressure frozen samples: the visibility of biological membranes is improved when the substitution medium contains water.
Biological membranes are often poorly visible with the electron microscope after high-pressure freezing and freeze-substitution. The water content of the sample and of the substitution medium is one factor among others that strongly influences membrane visibility. In order to investigate this effect, high-pressure frozen yeast cells, rat-pancreas tissue and arthropod tissue were freeze-substituted with and without adding water to the substitution medium. The visibility of the biological membranes was generally improved if the substitution medium contained 1-5% water. The effect was especially pronounced in yeast cells, where membrane visibility was poor after freeze-substitution with water-free medium but good after addition of 5% water to the substitution medium.
D000818:Animals; D002462:Cell Membrane / Q000648:ultrastructure*; D003445:Crustacea; D015925:Cryopreservation / Q000379:methods; D003451:Cryoprotective Agents; D004847:Epithelial Cells / Q000737:chemistry / Q000648:ultrastructure; D017110:Freeze Substitution / Q000379:methods*; D008854:Microscopy, Electron / Q000379:methods*; D010179:Pancreas / Q000737:chemistry / Q000648:ultrastructure*; D051381:Rats; D017208:Rats, Wistar; D012160:Retina / Q000737:chemistry / Q000648:ultrastructure*; D014021:Tissue Preservation; D014867:Water; D015003:Yeasts / Q000737:chemistry / Q000648:ultrastructure*
D003451:Cryoprotective Agents; D014867:Water