STAINING OF FLAGELLA. 117 



solution makes the fluid more permanent without impairing its pro- 

 perties. 



B. The Stain. To 100 c.c. of a filtered saturated solution of 

 aniline oil in water add I c.c. of a i per cent solution of sodium 

 hydrate. The aniline water is ordinarily neutral. The addition of 

 the soda renders it slightly alkaline. To this solution add 4-5 grm. 

 of solid methylene-violet, methylene-blue or fuchsin, and shake well. 

 When a preparation is to be stained, filter a few drops on to the 

 cover-glass. 



Make a film as above described, and holding the cover-glass in a 

 pair of forceps, pour on as much of the mordant A as the cover-glass 

 will hold. Heat it carefully above a flame till steam begins to rise 

 and then move the preparation gently in and out of the hot-air column 

 over the flame for about a minute. Wash well in distilled water till 

 every trace of mordant appears to be gone. If necessary, wash with 

 absolute alcohol till only the film itself appears tinted violet with the 

 mordant. Filter a few drops of stain B on to the cover, again heat 

 till steam rises and leave in the warm stain for one minute. Wash 

 well in distilled water, dry, and mount in xylol balsam. 



3. Van Ermengettfs Method for Staining Flagella. 



The films are prepared as above described. Three solutions are 

 here necessary : 



Solution A. (Bain fixateur) 



Osmic acid, 2 per cent solution . I part 



Tannin, 10-25 P er cent solution . . . .2 parts 



Place the films in this for one hour at room temperature, or heat 

 over a flame till steam rises and keep in the hot stain for five minutes. 

 Wash with distilled water, then with absolute alcohol for three to four 

 minutes, and again in distilled water, and treat with 



Solution B. (Bam sensibilisatenr) 



.5 per cent solution of nitrate of silver in distilled water. Allow 

 films to be in this a few seconds. Then without washing transfer to 



Solution C. (Bain reducteur et reinforcateur) 



Gallic acid . . . . . . 5 gr. 



Tannin . . ... . . . 3 gr. 



Fused potassium acetate . . . . 10 gr. 



Distilled water 350 c.c. 



Keep in this for a few seconds. Then treat again with Solution B 

 till the preparation begins to turn black. Wash, dry, and mount. 

 It is better, as Mervyn Gordon recommends, to leave the specimen 



