STAINING PROPERTIES 347 



alcohol, etc. If, therefore, a pathologic product containing tubercle 

 bacilli and other bacteria is treated with an intense hot staining solution 

 and then subjected to the action of a decolorizing fluid and afterward 

 counterstained with a contrast stain, the tubercle bacilli will have 

 kept the first stain, while other bacteria have been dyed with the 

 counterstain. In making use of this principle, tubercle bacilli can 

 easily be stained and distinguished from other bacteria. There are, 

 however, a few bacilli which act in a manner similar to the tubercle 

 organisms These and the methods for their differentiation will be 

 considered below. The bacilli of this group are known as acid-fast 

 bacilli. 



STAINING TUBERCLE BACILLI IN FLUID AND SEMIFLUID MATERIAL. 

 Reagents Required: 



(a) Ziehl's Carbol-fuchsin. 

 1 Carbolic acid, 5 c.c. 



2. Distilled water, 100 c.c. 



3. Basic fuchsin crystals, 1 gram. 



4. Absolute alcohol, 10 c.c. 

 Dissolve 1 in 2 and 3 in 4 and mix. 



(6) 10 per cent, watery solution of nitric acid. 



(c) 95 per cent, alcohol. 



(d) Ordinary watery solution of methylene blue. 

 Steps in Staining, Decolorizing, Counter staining. 



1. Prepare a cover-glass from suspected material, preferably 

 selecting some of the cheesy flocculi, as they are found in sputum, 

 pus, etc. After such material has been spread on a cover-glass, 

 allow it to become air dry, and fix by passing three times through a 

 flame. 



2. Apply to the cover-glass, held in a suitable forceps, enough of 

 Ziehl's carbol-fuchsin so that the whole surface is well covered; now 

 hold high over a small flame until the staining solution boils. Then 

 set it aside for a few minutes to permit the hot staining solution to 

 act well. 



3. Pour off the hot staining solution and wash the cover-glass in 

 ordinary tap water or in distilled water. 



4. Dip rapidly into the 10 per cent, watery nitric-acid solution, 

 and at once wash freely in 95 per cent, alcohol. Continue washing 

 the cover-glass, which is held in the forceps, in alcohol until apparently 

 all of the red stain has been washed out, i. e., until the cover-glass 

 again appears almost unstained. 



5. Dry between filter paper and for three or four minutes apply a 

 thin watery solution of methylene blue. 



6. Wash in water, dry between filter papers, mount on a slide in 

 the usual manner. 



Result of the Procedure Employed. The tubercle bacilli are stained 

 red; all other bacteria, yeast cells, etc., and cell nuclei are stained 

 blue. 



