THE STAINING OF BACTERIA. 47 



Besides the foregoing means, which will stain the bacteria 

 in films, bacteriologists adopt special methods for the staining 

 of bacteria in tissues, and for the staining of spores, flagella, 

 and the capsules of bacteria. 



V. The Staining of Capsules. 



1. Welch's Glacial Acetic Acid Method. 



Prepare the cover-glass in the usual way. Cover the film 

 with glacial acetic acid, pour off the acid immediately, do not 

 wash in water, cover the film with anilin-water gentian-violet 

 solution for three or four minutes, wash in a 0.5 to 2 percent, 

 solution of sodium chloride, dry, mount, and examine. 



The acetic acid coagulates the mucin of the capsules and 

 renders the same distinctly visible. 



2. Johne's Method. 



A cover-glass film prepared in the usual way is covered 

 with a solution of gentian- violet and heated until steam rises. 

 The stain is then washed off in water, and the cover-glass put 

 into a 2 per cent, acetic acid solution for from ten to fifteen 

 seconds. It is again washed in water, dried, and mounted in 

 balsam. 



VI. The Staining of Spores. 



Though with the ordinary method of staining, spores in 

 bacteria may be recognized by their highly refractive appear- 

 ance and by the fact that they have not taken the color, they 

 may be stained themselves, however, by special methods. 



The First Method (Abbott's). 



A cover-glass preparation is covered with Loeffler's alkaline 

 methylene-blue solution and held by its edge with forceps over 

 the Bunsen burner flame until the fluid begins to boil. It is 

 then removed from the flame, and after a few seconds heated 

 again. This step is repeated a number of times for one or 

 two minutes, after which it is washed in water, and decolor- 



