30 STAINING METHODS. 
three hundred parts); then washed in alcohol, dried, and, if for per- 
manent preservation, mounted in balsam. 
METHODS OF STAINING THE TUBERCLE BAcILLUS.—Numerous 
methods of staining the tubercle bacillus in sputum dried upon a 
cover glass have been proposed, but we shall only give here two or 
three of the most approved methods, either one of which may be 
relied upon for satisfactory results if carefully followed. 
1. The Ehrlich-Weigert Method.—Placein a watch glass a little 
of the aniline-methyl-violet solution (No. 2); float upon the surface 
of this the cover glass ‘with the dried film downward ; heat over a 
small flame until it begins to steam, then allow it to stand for from 
two to five minutes ; decolorize ina tray containing one part of nitric 
acid to three parts of water—the cover glass, held in forceps, is gently 
moved about’in the decolorizing solution for a few seconds. It is 
then washed off in sixty-per-cent alcohol to remove the remaining 
blue color—this usually takes but a second or two—and then in water. 
For a contrast stain a saturated aqueous solution of vesuvin may be 
used, a few drops being left upon the cover glass for five minutes. 
The stained preparation is then washed, dried, and mounted in 
balsam. 
2, The Ziehl-Neelson Method.—Float the cover glass upon the 
carbol-fuchsin solution (No. 3); heat gently until steam commences 
to rise—from three to five minutes’ time will usually be sufficient ; 
wash off in water, and decolorize in nitric or sulphuric acid, twenty- 
five-per-cent solution, then in sixty-per-cent alcohol for a very short 
time to remove remaining color from albuminous background; wash 
well in water and mount in Canada, balsam. 
3. Friedldnder’s Method.—Spread and dry the sputum upon 
the slide ; fix by passing the slide three times through the flame of 
an alcohol lamp or Bunsen burner ; place upon the dried film three or 
four drops of carbol-fuchsin (No. 3); heat gently over a flame until 
steam is given off ; wash in a dish of distilled water ; drain off excess 
of water, and adda few drops of the following decolorizing solution: 
Acid, nitric, pure, ‘ : : : 5 ce. 
Alcohol (eighty per cent), : ; “to 100 ce. 
—usually the preparation will be decolorized in about half a minute ; 
wash in water ; add a few drops of an aqueous solution of methylene 
blue as a contrast stain ; allow the stain to act for about five minutes, 
without heating ; wash again in water, dry, and mount in balsam, 
or for a temporary mount use a drop of cedar oil. 
4. Gabbett’s Method.—This is a slight modification only of a 
very useful method recommended by B. Frankel in 1884. The con- 
trast stain is added to the decolorizing solution. After staining witb 
