102 MICROSCOPIC METHODS. 



or fuchsin with aniline oil or carbolic acid or other mordant 

 will stain the bacilli named, but the following methods are 

 most commonly used : 



Ziehl-Neelsen Carbol- Fuchsin Stain. 



Basic fuchsin . . . . i part 



Absolute alcohol . . . .10 parts 

 Solution of carbolic acid (i : 20) . 100 parts 



1. Place the specimen in this fluid, and having heated it till steam 

 rises, allow it to remain there for five minutes, or allow it to remain in 

 the cold stain for from twelve to twenty-four hours. (Films and 

 paraffin sections are usually stained with hot stain, loose sections with 

 cold ; in hot stain the latter shrink.) 



2. Decolorise with 20 per cent solution of strong sulphuric acid, 

 nitric acid, or hydrochloric acid, in water. In this the tissues become 

 yellow. 



3. Wash well with water. The tissues thus regain a faint pink 

 tint. If the colour is distinctly red, the decolorisation is insufficient, 

 and the specimen must be returned to the acid. As a matter of prac- 

 tice, it is best to remove the preparation from the acid every few 

 seconds and wash in water, replacing the specimen in the acid and 

 re-washing till the proper pale pink tint is obtained. 



4. Contrast stain with a saturated watery solution of methylene-blue 

 for half a minute, or with saturated Bismarck-brown for from two to 

 three minutes. 



5. Wash well with water. Dehydrate, clear, mount. 



FraenkePs modification of the Ziehl-Neelsen Stain. 



Here the process is shortened by using a mixture 

 containing both the decolorising agent and the contrast 

 stain. 



The sections or films are stained with the carbol-fuchsin as above 

 described, and then placed in the following solution : 



Distilled water . . . . -5 parts 

 Absolute alcohol . . . 30 parts 



Nitric acid . . . . .20 parts 



Methylene-blue in crystals to saturation. 



They are treated with this till the red colour has quite disappeared and 

 been replaced by blue. 



