310 HISTOLOGICAL TECHNIQUE. 



10. Gram's Solution. — This is a differentiating agent used in con- 

 nection with anilin-gentian-violet : 



Iodine, 1 gram. 



Potassium iodide, 2 grams. 



Distilled water, 300 ec. 



The specimens are first overstained with the gentian-violet solu- 

 tion. They are then washed in water and placed in Gram's solution 

 for from three to five minutes. While in this solution they turn a 

 brown color, and the combination between the coloring-matter and 

 some of the elements of the specimen is loosened. The specimen 

 is then transferred to 95 per cent, alcohol, in which it remains until 

 no more color is given oif. If sufficient color has not been removed, 

 the treatment with Gram's solution and alcohol may be repeated. 

 After this differentiation the specimen may be dehydrated, cleared, 

 and mounted ; or a contrast-stain may be used before those manipu- 

 lations. This is a useful method for staining bacteria in sections 

 of tissue when the species of bacteria are such as resist the decolor- 

 izing action of the iodine. In this respect different species of bac- 

 teria differ greatly, and the method is commonly employed in bac- 

 teriological work to distinguish those species which retain the stain, 

 or are " positive to Gram," from those which are decolorized or 

 " negative to Gram." 



11. Van Giesen's Picric Acid and Acid Fuchsin Stain. — 



Aqueous solution of acid fuchsin, 1 per cent., 5 cc. 



Saturated aqueous solution of picric acid, 100 " 



This solution serves to stain fibrous intercellular substances. It 

 is used in the following manner : 



1. Slightly overstain with alum hematoxylin j e. <j., Bohmer's 

 hsematoxylin. 



2. Wash thoroughly in distilled water. 



3. Stain in Van Giesen's dye for five minutes. 



4. Wash in water. 



5. Dehydrate in 95 per cent, alcohol. 



6. Clear in oil of origanum. 



7. Mount in xylol-balsam or xylol-dammar. 



The tissues should have been fixed in a corrosive-sublimate solu- 

 tion or one containing eliminates ; e. g., Midler's fluid, Zenker's 



