Freeze‐drying of articular cartilage: Investigation of rat femoral heads by SEM

The results of comparative investigations of freeze‐drying of joint cartilage which had not been separated from the underlying bone are reported. Fixation and ethanol and amyl acetate substitution procedures cause marked loss of ground substance and thus create surface structures which are not present to the same degree in cartilage which is simply freeze‐dried. Indeed, it is questionable whether these structures are present in vivo. They were most highly developed in critical‐point‐dried cartilage. The main difficulty with freeze‐drying is the prevention of damage caused by ice crystals. A tissue temperature of −130°C should always be aimed for and, because of the rapid growth of ice crystals, the temperature should never be allowed to rise above −90°C. These conditions can be fulfilled if the specimen is in good thermal contact with the cold stage and if the cooling device is equipped with a condenser which is cooled with liquid nitrogen. The better the tissue was processed, the smoother was the resulting cartilage surface and the greater was the degree to which the chondrocytes fitted the walls of their lacunae. At tissue temperatures above −90°C, the first change which became apparent was destruction of the cell membranes. This was followed by almost complete elution of the ground substance from between the fibers and, finally, the cell nuclei were also destroyed. This damage caused by ice crystals exposed fibrous structures on the surface of the cartilage, and the appearance of these structures was superior to that produced by enzymatic methods of preparation. The disruption caused by freezing uncovered intracellular structures.