THE STAINING OF BACTERIA. 55 



The advantages of this method are that it is generally ap- 

 plicable. Bacteria are not robbed of their color, and the 

 tissue is sufficiently decolorized to render the bacteria visible 

 and to admit of a contrast-stain. 



5. Ziehl-Neelsen's Method. 



The sections are warmed in a solution of carbol-fuchsin for 

 one hour at a temperature of about 45 to 50 C., decolorized 

 for a few seconds in a 5 per cent, sulphuric acid solution, then 

 put into 70 per cent, alcohol, then in absolute alcohol for a few 

 seconds to dehydrate, then in xylol to clear, and mounted 

 on a slide in xylol balsam. 



6. Revised Directions for Making and Using the Wright 



Blood-stain. 



To a 0.5 per cent, aqueous solution of sodium bicarbonate 

 add methylene-blue (B. X. or "medicinally pure") in the 

 proportion of 1 gm. of the dye to each 100 c.c. of the solu- 

 tion. Heat the mixture in a steam sterilizer at 100 C. for 

 one full hour, counting the time after the sterilizer has be- 

 come thoroughly heated. The mixture is to be contained in 

 a flask or flasks of such size and shape that it forms a layer 

 not more than 6 cm. deep. After heating, allow the mixture 

 to cool, placing the flask in cold water if desired, and then 

 filter it to remove the precipitate which has formed in it. It 

 should, when cold, have a deep purple-red color when viewed 

 in a thin layer by transmitted yellowish artificial light. It 

 does not show this color while it is warm. 



To each 100 c.c. of the filtered mixture add 500 c.c. of a 

 0.1 per cent, aqueous solution of "yellowish, water-soluble " 

 eosin, and mix thoroughly. Collect the abundant precipitate 

 which immediately appears on the filter. When the pre- 

 cipitate is dry, dissolve it in methylic alcohol (Merck's 

 "reagent") in the proportion of 0.1 gm. to 60 c.c. of the 

 alcohol. In order to facilitate solution, the precipitate is to 

 be rubbed up with the alcohol in a porcelain dish or mortar 

 with a spatula or pestle. This alcoholic solution of the pre- 

 cipitate is the staining fluid. It should be kept in a well- 



