Dehydration for Embedding 25 



the tissues. Evaporation of water may be accomplished by several 

 methods or combinations of methods. The most practicable are as 

 follows: 



1. In an incubator oven at 35 to 40 °C. 



2. In a desiccator at room temperatures or in the above oven. 



3. In a vacuum desiccator or vacuum oven. 



If the glycerin solution becomes colored or turbid during evap- 

 oration it may be replaced with fresh glycerin solution of the same 

 concentration. When the volume of the solution has been reduced 

 by evaporation to one-half of the original volinne, the glycerin con- 

 centration is approximately 10 7c, and the liquid may be replaced 

 with fresh 10% glycerin and the evaporation continued. Most of the 

 water can be removed by evaporation, especially in vacuum. After 

 a nearly anhydrous condition is attained, the tissues are firm enough 

 to withstand transfer directly into anhydrous alcohol. Change the 

 alcohol at least twice, and proceed with the graded transfer to the 

 desired paraffin solvent as described below, or proceed with one of 

 the whole-mount methods (Chap. 10) . 



TRANSFER TO A SOLVENT OF PARAFFIN (CLEARING) 



After the use of dehydrating agents that are not solvents of 

 paraffin, the dehydrated tissues are transferred to a solvent. The term 

 clearing, applied to this transfer, is derived from the fact that some 

 paraffin solvents render the tissues transparent. The clearing action 

 is merely incidental to the function of the reagent, to serve as a 

 solvent of paraffin. The most common solvents are xylene (xylol) and 

 chloroform. Either reagent may be objectionable or even toxic to 

 some workers. Xylene is inexpensive and is by far the most widely 

 used solvent. Chloroform is more expensive, but it is less likely to 

 be toxic. Benzene and toluene can be used, but their lower boiling 

 points increase the fire hazard. 



As in the case of dehydration, a graded series is used for clearing. 

 After dehydration in ethyl alcohol, the following absolute alcohol- 

 xylene series is used. For critical cytological work 10 gradations have 

 been recommended. The interval in each mixture ranges from 1/2 hr. 



