MICROSCOPIC EXAMINATION AND STAINING METHODS 103 



Aqueous solution of gentian-violet: 



Saturated alcoholic solution of gentian-violet 2.5 c.c. 



Distilled water 47.5 c.c. 



Anilin gentian-violet (Ehrlich's) : 



Saturated alcoholic solution of gentian-violet 6 c.c. 



Absolute alcohol 5 c.c. 



Anilin water 50 c.c. 



Anilin water is prepared by adding 2 c.c. of anilin to 98 c.c. of 

 distilled water and shaking vigorously for several minutes. It 

 should then be filtered until clear. 



Carbol or phenol fuchsin (Ziehl's) : 



Saturated alcoholic solution of fuchsin 5c.c. 



Solution of phenol, 0.5 per cent 45 c.c. 



Bismarck-brown: This is commonly used as a saturated aqueous 

 solution. 



Gabbett's methylene-blue: 



Methylene-blue, dry 2 gm. 



Sulphuric acid 25 c.c. 



Distilled water 75 c.c. 



Preparation of a Stained Mount. A drop of water about the 

 size of a pinhead is placed upon a clean cover-glass. With a 

 sterile platinum needle remove a small portion of the material to 

 be examined and mix thoroughly in the drop of water. When the 

 bacteria are in bouillon or other liquid media, the drop of water 

 is unnecessary. This is then spread in a thin film over the surface 

 of the glass and dried. The film is next fixed by passing the 

 cover-glass, film up, through the flame of the Bunsen burner three 

 times. The stain is placed upon the glass and allowed to act for 

 a few seconds to ten minutes, depending upon the organism and 

 the stain used. This is then washed in water until no more stain 

 comes off. It is dried between filter-paper and placed film down 

 upon a drop of water on a slide and examined under the microscope. 

 If satisfactory, it may be floated off with water, dried, and placed 

 film down on a drop of Canada balsam on the slide. 



In many laboratories the use of the cover-glass is largely 

 dispensed with, and certainly routine examinations of many kinds 

 can be more conveniently made by means of films prepared 



