⑤ Dehydration

Completely remove water from the tissue through replacment with ethanol to enable the permeation of paraffin, which is immiscible with water, into the tissue.


  • Place tissue in 70% ethanol and then place in a shaker overnight.
  • Divide the tissue into large (e.g., cerebrum), medium (e.g., cerebellum), and small (e.g., brain stem, spinal cord) sizes.
  • Place two-ply filter paper on the bottom of a wide-mouthed container.
  • Place each different-sized tissue into containers filled with ethanol.
  • Place each tissue into a container and ensure that it is in contact with a flat surface to prevent deformation when shrinking occurs due to the dehydration process.
  • Place in a shaker or other suitable device.
  • Turn the tissue over several times a day to ensure that the tissue is dehydrated from both sides. When using 95% ethanol I for dehydration, place the tissue cut side down as this solution causes the tissue to significantly shrink.
  • Immerse the tissue in a series of ethanol solutions of increasing concenttation.

Dehydration timeframe

Guide for dehydration time

Dehydration process

Solution used for dehydrationTissue sizes
70% ethanol
80% ethanol
95% ethanol Ⅰ
95% ethanol Ⅱ
95% ethanol Ⅲ
95% ethanol Ⅳ
100% ethanol Ⅰ
100% ethanol Ⅱ
100% ethanol Ⅲ
100% ethanol Ⅳ

Important points concerning dehydration

Tissue size immediately after the cutting out procedure.

Tissue size after immersion in 100% ethanol. The tissue immersed in 100% ethanol is obviously shrunken compared with the tissue immediately after the cutting out procedure.