METHODS OF STAINING. 423 



When required for use, this solution should be diluted with dis- 

 tilled water to about one-tenth of its strength. It is a good stain for 

 bacteria, and may also be used for staining the nuclei of tissues 

 either by itself, or after using eosin as a diffuse stain. An aqueous 

 solution of eosin, 5 per cent., is used for this purpose, the sections 

 being stained for about five minutes. They are then washed to 

 remove the excess of eosin, and stained in the diluted methylene- 

 blue for about an hour. After this they are again washed and 

 treated with absolute alcohol, which discharges the excess of blue. 

 They are then cleared with xylol and mounted in dammar or 

 Canada balsam, dissolved in xylol. The preliminary staining with 

 eosin may be omitted, when a contrast- or counters.tain is not 

 required. 



7. Aqueous Methylene-blue. This is usually prepared at the time 

 when needed by mixing one part of a saturated solution of the ani- 

 1 in-color in 95 per cent, alcohol with nine parts of distilled water. 

 It is frequently employed as a general stain for bacteria. 



Other anilin-colors, such as fuchsin, gentian-violet, methyl-violet, 

 and Bismarck-brown, may be kept in concentrated alcoholic solu- 

 tion, to be diluted in a similar manner just before use. When these 

 solutions are used for staining sections or cover-glass preparations 

 the adherent dye is washed off with water, after which the intensity 

 of the stain is reduced by the use of alcohol, 95 per cent, or abso- 

 lute, which bleaches the portions of the specimen which retain the 

 color with the least tenacity. If the action of the alcohol be main- 

 tained for too long a time, the color may be discharged from all 

 parts of the specimen. The method of overstating a specimen, 

 and then discharging the color from those parts where it is not de- 

 sired, is a common one. The process of discharging the color is 

 called the " differentiation " of the stain, because it serves to dis- 

 tinguish those elements which hold the color strongly from those 

 which part with it easily. 



8. Carbol-fuchsin. 



Saturated alcoholic solution of fuchsin, 10 cc. 



Aqueous solution of carbolic acid crystals, 5 per cent., 90 cc. 



This solution should always be carefully filtered before use. 



9. Anilin-gentian-violet. A. Ehrlich's formula : 



Saturated alcoholic solution of gentian- violet, 1.5 cc. 



Freshly prepared anilin-water, 8.5 cc. 



