STAINING METHODS. 273 



glass preparation to remain for a quarter to half an hour in a dry 

 chamber at 180 to 200 C. In order to obtain simultaneously 

 beautiful spore-staining, and also staining of the rest of the body 

 of the bacteria, we can best use Ehrlich's aniline-water fuchsin 

 method. This reagent is prepared by first shaking up purified 

 aniline oil in excess with distilled water for about one minute. 

 We can use about 5 c.c. aniline oil in 100 c.c. water. Let this stand 

 for about five minutes, and then filter throughj a filter which has 

 been previously damped with distilled water. The filtrate must 

 be as clear as water, and it is recommended that it should always 

 be freshly prepared. We then add to it an alcoholic solution of 

 fuchsin until the fluid begins to become distinctly opalescent. 

 The Jdried cover-glass preparation, after passing the suggested 

 three times through the flame, is allowed to float for an hour upon 

 this aniline-water-fuchsin solution, made hot. The spores and 

 the protoplasm of the bacteria are then uniformly stained. It is 

 recommended, however, to now pick out the spores from the 

 cytoplasm by means of double staining ; for which purpose, after 

 use of the aniline-water-fuchsin, as above, a watery or dilute 

 alcoholic solution of methylene blue is specially suited. This 

 double staining is most certain if the superfluous aniline-water- 

 fuchsin is removed by laying the preparation for a few seconds or 

 minutes in absolute alcohol, or even in alcohol feebly acidulated 

 with hydrochloric acid (J per cent.), and the methylene blue then 

 used. In many cases instead of alcohol or acidulated alcohol 

 dilute mineral acids must be used in order to obtain quite sharp 

 figures. A successful double-staining with aniline-water-fuchsin 

 and methlylene blue shows the spores stained red, the rest of the 

 bacterialj body more or less dark blue. Instead of the aniline- 

 water method, quite the same results follow the use of Ziehl's 

 carbolic-fuchsin solution for spore staining. The solution is pre- 

 pared by dissolving 1 part fuchsin in 100 parts 5 per cent, carbolic 

 acid and 10 parts alcohol. Both solutions, Ehrlich's and Ziem"s> 

 but especially the [former, enjoy a very wide use for bacterial 

 staining in general. Ehrlich's solution can also be prepared with 

 gentian violet or with methyl violet, instead of with fuchsin. 

 It only keeps for at the most a few days, and should in general 

 not be used till twenty-four hours after preparation ; Ziehl's 

 solution on the other hand keeps. The staining capacity of 

 Ehrlich's solution is heightened if to 100 c.c. of the saturated 

 aniline water 1 c.c. of a 1 per cent, solution of caustic soda is 



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