36 A MANUAL OF BACTERIOLOGY 



Exercise 61. Special Stains 



1. Capsule stain (Welch). 



a. Prepare and fix the film in the usual manner. 



b. Flood the slide with 2 per cent acetic acid, leaving it in 

 contact for two minutes. This swells and fixes the capsule so 

 it will take the stain. 



c. Blow off the acetic acid by the aid of a pipette. 



d. Immerse in aniline-gentian-violet for five to thirty seconds. 



e. Wash in water, dry, and mount, 



2. Flagella stain. 



a. Make an agar streak of the organism to be stained. 



b. After eighteen to twenty -four hours, by means of the plati- 

 num needle remove a portion of the growth (being careful to 

 avoid the culture medium) to a large drop of tap water on a 

 clean cover glass. Allow this to stand five minutes rather than 

 spread, as there is less danger of breaking off the flagella. 



c. Spread carefully two or three loopfuls of this drop on 

 each of several clean cover glasses and dry at room temperature. 



d. Fix by passing through' the flame, the cover glass being 

 held in the hand. 



e. Flood the cover glass thus prepared with the following 

 solution (mordant) : liquor ferri sesquichloridi diluted with dis- 

 tilled water 1 : 20, one part ; saturated aqueous solution tannic 

 acid, three parts. This mixture improves with age but should be 

 filtered before using. Allow to act one minute. 



/. Wash the cover glass in water and dry by blotting it be- 

 tween strips of filter paper. 



g. Stain with hot carbol-fuchsin for about one minute. 

 h. Wash in water, dry, and mount in balsam. 



3. Spore stain. 



a. Prepare and fix film in usual way. 



b. Cover the film with hot carbol-fuchsin and hold above a 

 small flame or lay the cover glass in a watch glass full of stain 

 on a hot steam bath. Replace the stain as it evaporates. Keep 

 the film covered with hot carbol-fuchsin for twenty or twenty- 

 five minutes. (Both spores and bacteria are now stained.) 



