Scanning Microscopy


Specimen preparation methods are very important in scanning electron microscopy (SEM) of nerve tissues. In the present study, a t-butyl alcohol freeze-drying device was used to prepare cerebellar cortex of the human and that of the rat at 15°C and 160 mm Hg. This method has been previously used with success in the preparation of other tissues, such as pancreas and trachea. Relatively large specimens (about 10 mm x 15 mm x 1 mm) of formalin-fixed human and glutaraldehyde-Millonig buffer perfused (1 hour) Wistar rat were rinsed in water, dehydrated in a series of ethanols, immersed in t-butyl alcohol, and then placed in the new freeze-drying device. The specimens were cut with a razor, freeze-dried without acid or alkali digestion, mounted on stubs, and sputter-coated with gold. This new preparation method allowed a higher magnification examination of surfaces of cells and fibers of the human cerebellar cortex compared to the critical point drying method. This was valid for Purkinje cell bodies with axons, dendrites with climbing fibers and climbing fiber glomeruli, and stellate neuron cell processes connected to the Purkinje cell dendrites. Lugaro cell, basket cells with axons, Golgi II cell, mossy fiber glomerulus with granule cell dendrites, satellite Bergmann glial cells with processes, and many microtubule-like fibrous structures on the inside of Purkinje cell dendrites were observed. Furthermore with this method, the glutaraldehyde-Millonig buffer perfused cerebellar cortex of the Wistar rat shows better three-dimensional images than the formalin-fixed human cerebellar cortex.

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