Specimen preparation of the human cerebellar cortex for scanning electron microscopy using a t-butyl alcohol freeze-drying device.

Abstract

A freeze-drying device was applied to t-butyl alcohol substituted nerve cells, fibers and synaptic terminals. Ten percent formalin-fixed human cerebellar cortex specimens were postfixed in 2.5% glutaraldehyde in 0.1 M cacodylate buffer solution and were rinsed three times in 5% sucrose solution in the same buffer. After the postfixed specimens were dehydrated using a graded series of ethanols and then transferred into a graded series of t-butyl alcohols (freezing point 25.4 degrees C), the t-butyl alcohol substituted specimens were freeze-dried at 15 degrees C and at high vacuum (5 x 10(-2) Torr). The freeze-dried specimens were sputter coated with gold. Scanning electron microscopy revealed synaptic terminals on the surfaces of a Golgi cell and a small flat polygonal cell. Rough somatic surfaces of granule cells were also observed.