MICROSCOPIC STUDY AND STAINING 103 



Decolorize with ninety-seven per cent alcohol until no further traces 

 of the stain can be washed out of the preparation. This takes usually 

 thirty seconds to two minutes, according to thinness of preparation. 



Wash in water. 



Counterstain with an aqueous contrast stain, preferably Bismarck 

 brown. 1 



PALTAUF'S MODIFICATION OF GRAM'S STAIN. 2 The staining fluid as 

 prepared by this modification possesses the advantage of retaining its 

 staining power for a longer period than does the anilin-water gentian- 

 violet described in the original method. 



The staining fluid is prepared as follows: 



3-5 c.c. anilin oil are added to 

 90 c.c. distilled water and 

 7 c.c. absolute alcohol. 



This mixture is thoroughly shaken and filtered through a moist 

 filter paper until clear. Then add: 



Gruebler's gentian-violet 2 gm. 



The fluid should stand twenty-four hours, during which a precipi- 

 tate forms. This is filtered before use. 



This gentian-violet solution retains its staining power for from 4 to 6 

 weeks. It is good only when a metallic luster develops on the surface. 



It is used in the following way: Spreads on cover-slips or slides are 

 dried and fixed as usual. 



Then apply: 



Anilin-water gentian-violet (as above), three minutes. 



Gram's iodin solution, two minutes. 



Absolute alcohol (with stirring), thirty seconds 



Counterstain, without washing in water, in aqueous fuchsin or in 

 weak carbol-fuchsin. 



STERLING'S MODIFICATION OF GRAM'S METHOD. 2 c.c. anilin oil -f- 

 10 c.c. 95% alcohol. Shake and add 88 c.c. distilled water. 5 grams 

 gentian violet are ground in a mortar and the anilin solution added 

 slowly while grinding. Filter. This solution keeps, and stains in one- 

 half to one minute. 



1 To make up Bismarck brown solution, prepare a saturated aqueous solution of 

 the powdered dye by heating. Cool and filter. Dilute 1 to 10 with distilled water. 



2 Sh&rnosky, Proc. N. Y. Pathol. Soc., Oct., 1909, n. s., ix, 5. 



