FLAGELLA STAINING 103 



Flagella, being processes extending from the capsule, are, like 

 the latter, hard to demonstrate. They are not stained by the 

 common bacterial stains. In general a powerful stain mixed with 

 a strong mordant must be employed. Some methods appear 

 to be not so much a staining method in the ordinary sense but 

 either a precipitaing of the stain in the substance of the flagella 

 or else a decomposition of silver salts in the flagella substance. 

 To stain flagella, a young culture grown on agar must be employed ; 

 glycerine agar must never be used. A mass of the organism is 

 gently mixed with a drop of distilled water until a uniform emul- 

 sion is made. A dozen cover-slips carefully washed and cleaned 

 by alcohol are thoroughly flamed in order to remove the slightest 

 trace of grease. The watery emulsion of bacteria is then spread 

 over the cover-slips evenly and thinly. After they are dry the 

 bacteria are fixed by holding them for a minute just above the 

 apex of the flame with the fingers. The following methods may be 

 pursued : 



Pitfield's Method Modified by Muir. 



Two solutions are necessary for this method. 



A. Mordant. 



10 percent watery solution tannic acid '. . . 10 c.c. 



Corrosive sublimate saturated water solution 5 c.c. 



Carbol-fuchsin solution 5 c.c. 



This forms a dense precipitate which must be removed by the centrifuge, 

 or sedimentation, and the clear fluid, or mordant, is stored in a bottle. It 

 keeps for two weeks. 



B. Stain. 



Saturated watery solution of alum 10 c.c. 



Saturated alcoholic solution gentian violet 2 c.c. 



This keeps but two or three days. 



Flood the cover-slip with the mordant and gently steam for one 

 minute, then wash and dry thoroughly, pour the stain on and 

 steam for one minute more. Wash, dry and mount. 



This method yields very good results. 



