STAINS AND STAINING METHODS 99 



a peculiar staining reaction which may serve as an aid to 

 their identification. But when an organism is being 

 investigated, examination in the fresh and living condition 

 must never be omitted, for it is only thus that its motility 

 and life-history can be studied. Only general methods 

 are detailed here ; special ones will be given when they 

 are required. 



(1) LofHer's alkaline methylene blue : 



Saturated alcoholic solution of methylene blue . 30 c.c. 

 Solution of caustic potash, 0-01 per cent. . . 100 c.c. 



A very useful staining solution. Cultures should be quite fresh, 

 or the organisms do not stain well. When the organisms are 

 mixed with extraneous material, as in smears, or there is much 

 debris, this is one of the best staining solutions to employ. Methyl- 

 ene blue preparations are, however, not very permanent, and in 

 hot countries rapidly fade. Thionine blue is then preferable. (See 

 also p. 100.) 



Film specimens are stained for three to ten minutes, and 

 sections half to twenty-four hours. 



(2) Carbol-methylene blue (Kiihne) : 



Methylene blue . . . . . .1-5 grin. 



Absolute alcohol . . . . . 10 c.c. 



Five per cent, aqueous solution of carbolic acid . 100 c.c. 



A more intense staining solution than the former, and very 

 useful for sections, which are stained for from half to six hours. 



(3) Anilin gentian violet : 



Saturated alcoholic solution of gentian violet . 30 c.c. 

 Anilin water ....... 100 c.c. 



The anilin water is prepared by shaking 3 c.c. of anilin with 

 90 c.c. of distilled water, allowing the mixture to stand for a few 

 minutes, and filtering. 



This solution is a useful general stain for films, which are stained 

 for two or three minutes, and is employed in Gram's method of 

 staining. It does not keep well. 



Instead of anilin gentian violet, a carbol-gentian violet may be 

 used, and keeps much better than the foregoing (saturated alcoholic 



