SECTION ONE 



Note: Tissues which have been treated with a fixative such as 

 Miiller or Zenker, containing potass, dichromate, must be washed 

 from two to twelve hours in running water, depending upon the 

 size and the nature of the tissue, before being transferred to Diox- 

 ane. 



2. Transfer to a mixture of equal parts of paraffin wax and Diox- 

 ane for half to one hour in a paraffin embedding oven. 



3. Transfer to pure paraffin wax, allowing a somewhat longer 

 time for impregnation than for tissues cleared by the orthodox 

 method. 



Note: The Dioxane can be used several times provided the 

 calcium chloride is changed. 



ETHYLENE GLYCOL 



A solvent, inflammable when heated, burning with an intense, 

 almost invisible flame; which may also be employed as a de- 

 hydrating and diflFerentiating agent for the Sudan colours, in place 

 of acetone-alcohol, 70% or 50% alcohol; giving stable solutions, 

 without loss of stain from the lipid particles. 



Solution required 



Technique : 



1. Prepare the staining solution by heating and stirring about 

 0*75 gm. of the dye with 100 ml. pure anhydrous ethylene glycol 

 to 100° C. Filter the solution while it is still hot, and again after 

 it has been allowed to cool. 



2. Cut frozen sections, from formalin fixed material, and wash 

 them in water for about five minutes to remove excess formalin. 



3. Dehydrate the sections by agitating them gently with a camel 

 hair brush for three minutes in each of two changes of pure 

 anhydrous ethylene glycol. 



4. Transfer the sections to the staining solution for five to 

 seven minutes, agitating them gently at intervals. 



5. Differentiate by agitating the sections gently in 85% ethylene 

 glycol in water for two to five minutes, controlling by examining 

 under the microscope at intervals. 



37 



