CHAPTER 8 



Dehydrating and Clearing 



General Principles. Dehydration means "the removal of water from." 

 It is a necessary step in the preparation of specimens for microscopic ex- 

 amination because most of the media in which they will be mounted are 

 not miscible with water. The term " clearing " is rather misleading. It is 

 used in microscopy to describe the process of Removing alcoho l from de- 

 hydrated tissues. This process is necessary because the resinous media 

 used for mounting many specimens and waxes used for embedding before 

 cutting sections are no more miscible with alcohol than they are with 

 water. Many of the reagents used for clearing have a high index of refrac- 

 tion so that they make objects saturated with them appear more trans- 

 parent or clearer. Most of the reagents used for dehydrating and clearing 

 cause considerable shrinkage of tissues. This does not matter in the case 

 of animal tissues since the shrinkage is uniform and does not much distort 

 the appearance of the specimen. Shrinkage in plant tissues must, how- 

 ever, be avoided at all costs because the cellulose walls do not contract 

 as much as the cell contents. It follows that a carelessly dehydrated or 

 cleared plant tissue will show the contents of the cell pulled away from 

 the wall. 



Ethyl Alcohol. The commonest reagent used for dehydrating is ethyl 

 alcohol, which is available in most laboratories both as neutral grain 

 spirits (95 per cent alcohol) and absolute alcohol (100 per cent alcohol). 

 The removal of the last 5 per cent of water from neutral grain spirits is 

 a very expensive operation, so that 95 per cent alcohol should be used 

 wherever possible. 



Were the majority of specimens merely to be thrown in 95 per cent 

 alcohol, the violent diffusion currents that would be set up would result 

 in the collapse of cavities or in the distortion of the specimen. It is cus- 

 tomary, therefore, to use these alcohols as a graded series. It is conven- 

 tional today to employ the series of 30 per cent, 50 per cent, 70 per cent, 

 90 per cent, and 95 per cent and to pass the specimen from one of these 

 strengths to the next, leaving it in each sufficiently long to become im- 



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