THE TUBERCLE BACILLUS 481 



acids. For this reason, this bacillus, together with some other bacteria 

 to be mentioned later, is spoken of as "acid-fast." The acid-fast nature 

 of the bacillus seems to depend upon the fatty substances contained in 

 it, 1 and has furnished the basis for differential staining methods. All 

 the staining methods devised for the recognition of the tubercle bacillus 

 thus depend upon the use of an intensely penetrating staining solution, 

 followed by vigorous decolorization which deprives all but the acid-fast 

 group of their color. Counterstains of any of the weaker dyes may 

 then be used to stain the decolorized elements. One of the first of the 

 staining solutions to be of practical use was the anilin-water-gentian- 

 violet solution of Ehrlich 2 (11 c.c. saturated alcoholic gentian- violet 

 to 89 c.c. 5 per cent anilin water). This dye, although of sufficient 

 penetrating power, has the disadvantage of deteriorating rapidly and 

 has in practice been almost entirely displaced by Ziehl's 3 carbol-fuchsin 

 solution. (Fuchsin 1 gm. in 10 c.c. alcohol absolute, added to 90 c.c. 

 5 per cent carbolic.) This staining solution is the one now in 

 general use and is employed as follows: Thin smears, on slides 

 or cover-slips, are covered with the dye and gently heated. In 

 the case of cover-glasses, these may be floated, face downward, 

 in staining x dishes filled with the dye. The dye is allowed to act 

 for about three minutes, steaming but not allowed to boil. At the 

 end of this time the preparation is washed either with 5 per cent 

 nitric acid, 5 to 20 per cent sulphuric acid, or 1 per cent hydro- 

 chloric acid, until most of the .red color has disappeared (a few 

 seconds), and the preparation appears pale pink. This results in 

 decolorization of all microorganisms with the exception of members 

 of the acid-fast group. - Thorough washing in 80 to 95 per cent alcohol 

 is now employed to complete the decolorization. The preparation 

 is then, rinsed in water and counterstained with 1 per cent aqueous 

 methy lene-blue . 



Tubercle-bacillus staining has been further simplified by Gabbett, 4 

 who combines decolorization and counterstaining. In this method 

 preparations are stained with Ziehl's carbol-fuchsin as in the preceding; 

 they are then rinsed in water and covered with a solution containing 

 methylene-blue 1 gram, concentrated sulphuric acid 25 grams, and 

 distilled water 100 c.c. This is allowed to act for from two to four 



1 Bienstock, Fort. d. Med., 1886; Weyl, Deut. med. Woch., 1891. 

 Ehrlich, Deut. med. Woch., 1882; Weigert, Deut. med. Wochi, 1885. 

 Ziehl, Deut. med. Woch., 1883; Neelsen, " Lehrb. d. allg. Path.," 1894 

 Gabbett, Lancet, 1887. 



