EXAMINATION AND STAINING OF BACTERIA 47 



on the other hand a distinctly red color remains the process should 

 be repeated until the proper tint is obtained. The slide is next 

 immersed in a 10 per cent watery solution of methylene blue, 

 washed in water, and dried. 



The above method is used for the group of organisms known 

 as " acid-fast," amongst which are the tubercle bacillus, the 

 leprosy bacillus, the smegma bacillus, the hay bacillus, and a 

 number of others. These organisms require a powerful dye con- 

 taining a mordant, and the staining process must be continued 

 a long time or its action aided by the application of heat. When 

 once stained, however, they resist the decolorizing action of 

 strong acids ; for this reason they are spoken of as " acid-fast." 

 Stained by the above method they appear under the microscope 

 to be bright red while any other organisms present take the 

 counterstain and appear blue. 



Spore Stain. Moeller's Method. The films are immersed 

 in chloroform for two minutes, washed in water, then covered with 

 5 per cent chromic acid one minute and again washed in water. 

 They are next placed in carbol fuchsin and the solution is heated 

 until it commences to steam. After remaining in the hot solution 

 for three minutes the slides are removed, washed in water, and 

 decolorized in 5 per cent sulphuric acid for five to ten seconds, 

 then washed in water and counterstained in 10 per cent aqueous 

 methylene blue one minute. By this method the spores appear red 

 while the remainder of the bacterial cell is stained blue. If bacilli 

 containing spores are stained with a watery solution of any of 

 the aniline dyes the spores remain unstained and appear as clear 

 spaces surrounded by stained protoplasms. 



Capsule Stain. Hiss' Method. Films are made in the usual 

 way and fixed by heat. The slide is then covered with a 5 per 

 cent watery solution of fuchsin or gentian violet and heated over 

 a Bunsen flame until it steams. The dye is washed off with a 

 20 per cent aqueous copper sulphate solution, after which it is 

 dried between layers of filter paper without further washing in 

 water. By this method the capsule appears as a faint blue halo 

 surrounding a dark purple or red cell body. 



