SECTION THREE 



SCARLET R or SUDAN 3 - ETHYLENE GLYCOL 



An improved technique for staining fat in plant tissues, offering 

 the following advantages : (a) The use of ethyl alcohol is obviated 

 and sections remain pliable and unshrunken; {b) excellent dif- 

 ferentiation without loss of stain from the fat; (c) More intense 

 staining of fat. 



Solutions required: 



A. Scarlet R or Sudan 3 . . . . i gm. 

 Ethylene glycol, pure, anhydrous.. 100 ml. 



Heat the ethylene glycol to 100-110° C. on a hot 

 plate, or in an oven for preference, but if these are not 

 available the bunsen flame will serve the purpose so 

 long as care is taken to ensure that the ethylene glycol 

 does not take fire. Stir in the dye until all or most of it 

 is dissolved; then cool and filter. 



B. Ethylene glycol . . . . • • 85 ml. 

 Distilled water . . . . . . 15 ml. 



C. Delafield or Ehrlich haematoxylin. 



Technique: 



1. Fix tissues in 10% formalin. 



2. Wash out fixative with water. 



3. Dehydrate the sections by agitating gently in the pure 

 ethylene glycol, anhydrous, for three to five minutes. 



4. Immerse the sections in the staining solution (Scarlet R or 

 Sudan 3) and agitate for two to five minutes. 



5. Differentiate by agitating gently, at intervals, from one to 

 five minutes in 85% ethylene glycol (solution B) controlling by 

 examination under the microscope while the specimen is still wet. 



6. Transfer the sections to distilled water and leave therein for 

 three to five minutes. 



7. Counterstain with Delafield or Ehrlich haematoxylin. 



8. Wash well in tap water. 



9. Mount in glycerine jelly or glycerine. 



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