28 Botanical Microtechnique 



Make three changes of anhydrous tertiar\ buiNl alc(jhol and pro- 

 ceed with infiltration in wax. 



An unfa\orable factor in the use of tertiary butyl alcohol is that 

 it solidifies at 25.5°C., a temperature that is not uncommonly attained 

 in laboratories and stock rooms. Provisions must be made to keep 

 this reagent fluid for immediate use. The low boiling point of 82.8°C. 

 presents some fire hazard. 



The butyl alcohols have greatly extended the range of usefulness 

 of the paraffin method by making it possible to cut materials that 

 are rendered hard and brittle by ethyl or propyl alcohol or acetone. 



THE DIOXAN METHOD 



Dioxan, diethylqne dioxide, is becoming widely accepted as a 

 dehydrating agent and paraffin solvent in the embedding of plant 

 materials. This reagent is miscible with water and may therefore be 

 progressively substituted for water in the tissues. Unlike the vigorous 

 dehydrating action of the alcohols or acetone, the substitution of 

 water by dioxan rs not associated with great plasmolyzing stresses. This 

 fact permits dehydration by rapid substitution. Tissues do not become 

 excessively brittle, and the histological details obtainable are equal 

 to those obtained by other methods. The dioxan method requires 

 nuich fewer separate operations than does any other method, and 

 the operations may be at widely spaced intervals, thus reducing the 

 burdensome routine of handling the specimens many times at frequent 

 intervals. 



Kill the material in the desired formula. After the optimum 

 fixing interval, wash in water if required by the formula. Animal 

 tissues are said to be transferable directly from the wash water into 

 pure dioxan, but plant cells are plasmolyzed by such treatment. 



Materials that were washed in water are transferred through the 

 following three grades at 4- to 12-hr. inlerxals. Wide latitude in these 

 intervals is permissible. 



(1) i/( dioxan 

 % water 



(2) 2/^ dioxan 

 1/^ water 



(3) Anhydrous dioxan. Replace the cork with a jierfec tlv drv one. 



Make two more changes of anhydrous dioxan after intervals of 

 4 to 8 hr. Proceed with progressive infiltration in paraffin as described 

 in the next chapter. 



Materials that were killed in FA A, or in anv fluitl tiiat is followed 



