42 BACTERIOLOGY 



which has previously been fixed in absolute methyl alcohol, is 

 floated on this mixture for about ten minutes. Considerable 

 practice is necessary before the best results are obtainable. 

 The method is especially useful in staining blood films, and 

 protozoa in blood, in feces or in culture. 



Leishman's Stain. Leishman has utilized the principle of 

 Jenner's stain 1 and has added to it the important additional 

 constituents found in polychrome methylene blue by substituting 

 this for the ordinary methylene blue used by Jenner. 



Solution A. To a i per cent solution of medicinally pure 

 methylene blue in distilled water add 0.5 per cent sodium car- 

 bonate and heat at 65 C. for 12 hours, then allow it to stand 10 

 days at room temperature. 



Solution B. Eosin extra B. A. (Griibler) o.i per cent solution 

 in distilled water. 



Mix Solutions A and B in equal amounts and allow to stand 

 six to twelve hours, stirring at intervals. Filter and wash the 

 precipitate thoroughly. Collect, dry and powder it. 0.15 gram 

 is dissolved in 100 c.c. of pure methyl alcohol to form the staining 

 solution. It keeps perfectly for at least five months. To stain, 

 cover the dried but unfixed film of blood with the staining solu- 

 tion. After 30 to 60 seconds add about an equal amount of 

 distilled water. Allow this mixture to act for five minutes. 

 Wash in distilled water for about one minute, examining the 

 specimen mounted in water under the microscope. Blot, dry 

 thoroughly, mount in balsam, or preserve the specimen as an 

 unmounted film. 



Numerous imitations or modifications of Leishman's stain 

 have been described. 



Giemsa's Stain. This stain contains certain of the essential 

 constituents of polychrome methylene blue and eosin, the whole 

 being dissolved in a mixture of glycerin and methyl alcohol. 

 Giemsa's Azur I is the substance methylene azure and his Azur 



1 Jenner (Lancet, 1899, I, p. 370) first employed the solution of eosin and methy- 

 lene blue in methyl alcohol as a stain for blood films. 



