728 PATHOGENIC PROTOZOA 



slides, covered with a glass and ringed with warm vaseline to prevent 

 evaporation. The preparation, to be of value, must be thin, and the 

 bloody mucus may be diluted with salt solution if necessary. Except 

 in hot weather, the slide should be examined on a warm stage, or the 

 slide may be warmed by placing heated coins on it, near the cover 

 glass. At least half a dozen slides should be examined before reporting 

 a negative result. 



Stained preparations are not difficult to prepare, although the 

 process requires some time and care. As in most zoological work, wet, 

 rather than dry, fixation is used. Thin smears are made on cover 

 glasses or slides and before they can dry are covered with or immersed 

 in Schaudinn's fluid. This is a mixture of two parts of a saturated 

 solution of bichlorid of mercury in normal salt solution and one part 

 of absolute alcohol. The mercuric solution is prepared by adding to 

 boiling normal salt solution a little more mercury than will dissolve; 

 on cooling, some of the bichlorid crystallizes out. At no stage of the 

 process must the preparation become dry or the smear is worthless. 



1. Fix in hot (60 C.) Schaudinn's fluid, 5 to 10 minutes. 



2. Harden in 70 per cent alcohol 10 to 30 minutes, then wash in 

 70 per cent alcohol to which a few drops of tincture of iodin have been 

 added until it is distinctly colored 10 minutes ; store in 70 or 80 per 

 cent alcohol until ready to stain. 



3. The Rosenbusch hematoxylin is quite satisfactory. Transfer the 

 slides to distilled water and change several times until they are free 

 from alcohol, then immerse in 3.5 per cent iron-alum solution for from 

 half an hour to over night. 



4. Stain in the following solution, after rapid washing in distilled 

 water : 



(a) 1 per cent hematoxylin in 95 per cent alcohol. 



(&) Saturated aqueous solution of lithium carbonate. 



Solution (6) is added to solution (a) until the mixture is a 

 cherry red, four or five drops of lithium to 10 c.c. of hematoxylin 

 is sufficient. 



The solution is either pipetted onto the slides or .they are im- 

 mersed in it. Stain from 20 minutes to over night. 



5. Wash thoroughly in distilled water. 



6. Differentiate with a weak iron-alum solution (three parts of 

 distilled water to one of the iron-alum solution is satisfactory), until 



