SECTION THREE 



SAFRANIN - ANILINE BLUE 



For plant tissues; particularly suitable for gymnosperm 

 ovules, archegonia, embryos and angiosperm stems and 



roots 



Solutions required: 



A. Safranin O, 2% in Cellosolve . . 100 ml. 

 Absolute alcohol . . . . . . 50 ml. 



Sodium acetate 4% aqueous . . 50 ml. 



Formaldehyde 40% . . . . . . 8 ml. 



B. Picric acid 0*5% in 95% alcohol. 



C. Ammonia solution (sp. gr. 0.88) . . 0-25 ml. 

 Absolute alcohol . . . . . . 100 ml. 



D. Aniline Blue, alcohol soluble, saturated in 

 equal volumes of cellosolve and absolute 

 alcohol. 



Technique: 



1. Take sections through to 70% alcohol in the usual manner. 



2. Stain from two to forty-eight hours, according to the nature 

 of the material, in solution A. 



Note: Gymnosperms require the minimum staining time, 



3. Wash thoroughly with running water to remove the excess 

 stain. 



4. Dehydrate and differentiate carefully with solution B. 



5. Immerse the preparation in solution C for half to one minute. 



6. Dehydrate morphological material in 95% alcohol, or cyto- 

 logical preparations in absolute alcohol. 



7. Counterstain for about one minute in a mixture consisting 

 of equal parts of solution D and clove oil. 



8. Clear in methyl salicylate and mount in Cristalite, Canada 

 balsam or Emexel mountant. 



Results: 



Lignified and cutinized cell walls, nuclei and chromosomes, 

 bright red. Cellulose cell walls and cytoplasm, blue. 



273 



