106 STAINING METHODS 



3. Wash thoroughly in water. 



4. Stain in haematoxylin solution (see page 95) for 

 thirty minutes. 



5. Wash in water. 



6. Differentiate in ferric ammonia sulphate solu- 

 tion for iJ-2 minutes, examining wet under micro- 

 scope during the process. 



To Stain Spores. 

 1. Single Stain. 



1. Prepare cover-slip film in the usual way. 



2. In fixing, pass the cover-slip film fifteen or thirty 

 times through the flame instead of only three. This 

 destroys the resisting power of the spore membrane 

 and allows the stain to reach the interior. 



3. Stain in the usual way with methylene-blue or 

 fuchsin. 



4. Wash in water. 



5. Dry and mount. 



2. Double Stain. 



1. Prepare and fix film in the usual way i. e., pass 

 three times through flame to fix. 



2. Cover the film with hot carbol-fuchsin and hold 

 in the forceps above a small flame until the fluid begins 

 to steam. Set the cover-slip down and allow it to 

 cool. Repeat the process when the stain ceases to 

 steam and continue to repeat until the stain has been 

 in contact with the film for twenty minutes. (This 

 stains both spores and bacteria.) 



3. Wash in water. 



4. Decolourise in alcohol, 2 parts; acetic acid, i per 

 cent., i part. (This removes the stain from everything 

 but the spores.) 



5. Wash in water. 



6. Mount the cover-slip in water and examine micro- 

 scopically with the J-inch objective. (Spores should 



