Staining 719 



remembered that the quantity of material examined by the 

 staining method is extremely small, so that a few bacilli in 

 a relatively large quantity of matter can easily escape dis- 

 covery. 



Staining the Bacillus in Sections of Tissue. Ehrlich's 

 Method for Sections. Ehrlich's method must be recom- 

 mended as the most certain and best. The sections of 

 tissue, embedded in paraffin, should be cemented to the slide 

 and then freed from the embedding material. They are then 

 placed in the stain for from twelve to twenty-four hours and 

 kept at a temperature of 37 C. Upon removal they are 

 allowed to lie in water for about ten minutes. The washing 

 in nitric acid (20 per cent.) which follows may have to be 

 continued for as long as two minutes. Thorough washing in 

 60 per cent, alcohol follows, after which the sections can be 

 counterstained, washed, dehydrated in 96 per cent, and abso- 

 lute alcohol, cleared in xylol, and mounted in Canada balsam. 

 Unna's Method for Sections. Unna's method is as follows : 

 The sections are placed in a dish of twenty-four-hour-old, 

 newly filtered Ehrlich's solution, and allowed to remain 

 twelve to twenty-four hours at the room temperature 

 or one to two hours in the incubator. From the stain 

 they are placed in water, where they remain for about 

 ten minutes to wash. They are then immersed in acid 

 (20 per cent, nitric acid) for about two minutes, and be- 

 come greenish black. From the acid they are placed 

 in absolute alcohol and gently moved to and fro until the 

 pale-blue color returns. They are then washed in three 

 or four changes of clean water until they become almost color- 

 less, and then removed to the slide by means of a section- 

 lifter. The water is absorbed with filter-paper, and then the 

 slide is heated over a Bunsen burner until the section be- 

 comes shining, when it receives a drop of xylol balsam and 

 a cover-glass. 



It is said that sections stained in this manner do not 

 fade so quickly as those stained by Ehrlich's method. 



Gram's Method. The tubercle bacillus stains well by 

 Gram's method and by Weigert's modification of it, but 

 these methods are ill adapted for differentiation. They 

 should not be neglected when no tubercle bacilli are demon- 

 strable by the other methods, as they are particularly well 

 adapted to the demonstration of such of the organisms as 

 may not be acid-proof. 



