82 PATHOGENIC BACTERIA. 



tion of the color. For such forms a more intense stain 

 must be employed. One of the best of these stains, 

 which can be employed by the given method both for 

 cover-glasses and tissues, is Loffler's alkaline methylene 

 blue: 



Saturated alcoholic solution of methylene blue, 30 ; 



i : 10,000 aqueous solution of caustic potash, 100. 



Some bacteria, as the typhoid-fever bacillus, decolorize 

 so rapidly as to contraindicate the use of acid for the dif- 

 ferentiation, washing in water or alcohol being sufficient. 

 Gram's Method of Staining Bacteria in Tissue. 

 Gram was the fortunate discoverer of a method of stain- 

 ing bacteria in such a manner as to saturate them with 

 an insoluble color. It will be seen at a glance what a 

 marked improvement this is on the method given above, 

 for now the stained tissue can be washed thoroughly in 

 either water or alcohol until its cells are colorless, with- 

 out fear that the bacteria will be decolorized. Its prose- 

 cution is as follows : The section is stained from five to 

 ten minutes in a solution of a basic anilin dye pure 

 anilin (anilin oil) and water. This solution, first devised 

 by Ehrlich, is known as Ehrlich's solution. The ordinary 

 method of preparing it is to mix the following : 



Pure anilin, 4 ; 



Saturated alcoholic solution of gentian violet, n ; 



Water, 100. 



Instead of gentian violet, methyl violet, fuchsin, or any 

 basic anilin color may be used. The mixture does not 

 keep well in fact, seldom longer than six to eight weeks, 

 sometimes not more than two or three ; therefore it is 

 best to prepare it in very small quantity by pouring 

 about i c.cm. of pure anilin into a test-tube, filling 

 the tube about one-half with distilled water, shaking 

 the mixture well, then filtering as much as is desired 

 into a small dish. To this the saturated alcoholic solu- 

 tion of the basic dye is added until the surface becomes 

 distinctly metallic in appearance. 



