SOLUTIONS AND STAINS' 29 



of methylene blue. The end-reaction is reached when enough 

 eosin has been added to neutralize all of the methylene blue 

 and its oxidation products. To determine this end-reaction, 

 place a drop of the mixture on a piece of filter paper, a slight 

 eosin halo appears around the drop, due to a slight excess of 

 eosin. As the precipitate is soluble in eosin, add only enough 

 excess of eosin to get the slight halo. Collect this precipitate 

 on a filter paper and dry in the incubator at 38 C. When 

 thoroughly dry, dissolve 0.06 gram in 20 c.c. of pure methyl 

 alcohol (acetone-free). This is the stock solution. For 

 use, filter the 20 c.c. and add to the filtrate 5 c.c. of methyl 

 alcohol. 



The dry powder kteps well: the alcoholic solution does not 

 keep well. Therefore it is better to make only enough of the 

 solution to last a couple of months. 



Method of Staining. i. Make films 'and dry in the air. 

 The film must dry quickly and must be protected from dust 

 and dirt. 



2. Cover the dry film preparation with the stain for one 

 minute (to fix). 



3. Add distilled water to the stain on the preparation, 

 drop by drop, until a yellow metallic scum begins to form 

 (to stain). Add the drops of water rapidly in order to pre- 

 vent precipitates on the stained film; practically add i drop 

 of water for every drop of stain used. Allow to stain for 

 five to ten minutes. 



4. Wash off the stain with distilled water. 



5. Wash in distilled water until the film has a pinkish tint. 



6. Blot dry with filter paper. Do not put on a cover- 

 glass or mount in liquid petroleum. 



Red cells are orange to pink; nuclei, various shades of violet; 

 eosinophile granules are red; neutrophile granules are yellow 

 to lilac; blood plates are purplish; malaria parasites: cyto- 

 plasm is blue and chromatin is metallic red to rose pink. 



Caution. Never heat a preparation that is to be stained 



