Staining Protozoa in Tissue 199 



STAINING PROTOZOA IN TISSUE. 



For this purpose the sections should be embedded in paraf- 

 fin, cut very thin, and cemented to the slides. 



Ordinary staining with hematoxylin and eosin is rarely of 

 much use. Methylene-blue and eosin is better, but still 

 more useful are the Romanowsky methods, and both the 

 Wright stain and the Marino stain can, with some modifica- 

 tion of the time of staining and washing, be employed with 

 good results. 



Still better and more satisfactory for certain protozoa are 

 the iron-hematoxylin and the Biondi stain. 



Heidenhain's Iron-hematoxylin.* Fix the tissue, by preference, in 

 Zenker's solution, though alcohol fixation will do. Embed in 

 paraffin, cut very thin, and fix to the slide. 



1. Stain from three to twelve hours in 2.5 per cent, solution of 

 violet iron-alum (sulphate of iron and ammonium). The sec- 

 tions should be stood vertically in the solution, so that no pre- 

 cipitate may form upon them. 



2. Wash quickly in water. 



3. Stain in a 0.5 per cent, ripened alcoholic solution of hematoxylin 

 for from twelve to thirty-six hours. 



4. Wash in water. 



5. Differentiate in the iron-alum solution, controlling the results 

 under the microscope. The section should be well washed in a 

 large dish of tap-water before each examination to stop decoloriza- 

 tion. 



6. Wash in running water for a quarter of an hour. 



7. Pass through alcohol, xylol, and mount in xylol balsam. 



A counterstain with Bordeau R. before or with rubin S. after the 

 iron stain is sometimes useful. 



Biondi-Heidenhain Stain* The tissues must be fixed in Zenker's 

 or corrosive sublimate solutions. Embed in paraffin, cut very 

 thin, fix to the slide. 



Stain I. Orange G 8 grams. 



Water 100 



II. Acid fuchsin \ 



or Rubin S. J 



Water 100 



III. Methyl-green 8 grams. 



Water 100 



Let the solutions stand for several days, occasionally shaking 

 the bottles to make sure that a saturated solution of each is 

 secured. At the end of the time set, mix the solutions in the 

 following preparations : 



1 100 parts. 



II 20 



III 50 



* Mallory and Wright, "Pathological Technique," 1911, p. 309- 

 f Modified from Mallory and Wright, " Pathological Technique," 

 1911, p. 289. 



