378 BACILLI IN CHRONIC INFECTIOUS DISEASES. 



gently heated at intervals for a minute or two. The cover glass is 

 then washed in water, and the film will be seen to have a uniform 

 deep-red color. The next step consists in decolorization in the acid 

 solution (twenty-five-per-cent solution of nitric or of sulphuric acid). 

 The cover glass is gently moved about in this solution for a few 

 seconds, and the color will be seen to quickly fade to a greenish 

 tint. The object is to remove all color from the cells and the al- 

 buminous background, so that the bacilli, which retain their color in 

 presence of the acid, may be clearly seen. The preparation is next 

 washed in dilute alcohol (sixty per cent) to remove the fuchsin 

 which has been set free by the acid. If decolorization was not car- 

 ried far enough the film will be seen to still have a red color, espe- 

 cially in places where it is thickest, when it is removed from the 

 dilute alcohol and washed out in water. In this case it will be 

 necessary to return it to the acid solution and again wash it in the 

 dilute alcohol and in water. It may now be placed in a solution 

 of methylene blue or of vesuvin for a contrast stain. The tubercle 

 bacilli are distinguished by the fact that they retain the red color 

 imparted to them in the fuchsin solution, while other bacteria pre- 

 sent, having been decolorized in the acid solution, take the contrast 

 stain and appear blue or brown, according to the color used. The 

 double-stained preparation, after a final washing in water, may be 

 examined at once, or dried and mounted in balsam for permanent 

 preservation. 



Of the various other methods which have been proposed, that of 

 Frankel, as modified by Gabbett, appears to be the most useful. This 

 consists in staining as above directed with Ziehl's carbol-f uchsin solu- 

 tion, and in then placing the cover glass directly in a second solution 

 which contains both the acid for decolorizing and the contrast stain. 

 This second solution contains twenty parts of nitric acid, thirty parts 

 of alcohol, fifty parts of water, and sufficient methylene blue to make 

 a saturated solution (one to two parts in one hundred). After re- 

 maining in this solution for a minute or two the cover glass is washed 

 in water, and upon microscopical examination the tubercle bacilli, if 

 present, will be seen as red rods which strongly contrast with the 

 blue background. 



The methods recommended for cover-glass preparations may also 

 be used for staining the tubercle bacillus in thin sections of tuber- 

 culous tissues, except that it is best not to employ heat. The sec- 

 tions may be left for an hour in the carbol-fuchsin solution, or for 

 twelve hours in the Ehrlich-Weigert tubercle stain eleven cubic 

 centimetres of saturated alcoholic solution of methyl violet, ten cubic 

 centimetres of absolute alcohol, one hundred cubic centimetres of ani- 

 line water. They should then be decolorized by placing them for 



