STAINING TREPONEMATA 47 



Mallory's Iron Haematoxylin Method 



I have obtained beautiful staining with this simple method. The great point in 

 technic is the watching of the differentiation. 



Treat sections or moist smears fixed by Giemsa's method with 10% aqueous solu- 

 tion of ferric chloride for three to five minutes. Then drain off iron solution, blot 

 the section and stain for four minutes with a freshly made solution of i % haematoxylin 

 in water. To make this add a few small crystals to 4 or 5 c.c. water in a test-tube 

 and dissolve by heat. The stain deteriorates after twenty-four hours. Wash in 

 water. Differentiate with a Y% aqueous solution of ferric chloride. The dif- 

 ferentiation is complete in from a few seconds to one or more minutes according to 

 depth of staining and thickness of film. Wash in water, pass through alcohols and 

 xylol and mount. 



Spirochaete Staining Method (Fontana) 



The smears must be air dried, not fixed by heat. Cover films with Huge's fluid 

 which is i c.c. acetic acid, 20 c.c. formalin and 100 c.c. distilled water. Flood films 

 several times with this fluid. Wash in water and cover with a mordant of 5% tannic 

 acid in i % carbolic acid solution. Heat the mordant on the slide until steam arises, 

 and allow the heated mordant to act about thirty seconds. Wash in water and 

 without drying pour on the silver stain, heat until steam arises, leave heated stain 

 on for thirty seconds, wash in water, blot, dry and examine with immersion objective. 

 The treponemata are brownish to black. 



To make the silver stain make a J4% solution of silver nitrate in distilled water. 

 Then add drop by drop ammonia until a slight turbidity is produced; only a trace 

 of ammonia is required and any excess again clears up the solution and makes it 

 useless. 



