184 MICROMETRY AND BLOOD PREPARATIONS 



tilled water. If i to 1000 potassium carbonate solution is used instead 

 of water it stains more deeply. 



The alkaline diluent is used to obtain the course stippling in malig- 

 nant tertian (Maurer's clefts). Having fixed the smear with methyl 

 alcohol for one to five minutes, pour on the diluted stain, and after 

 fifteen to thirty minutes wash off and continue washing with distilled 

 water until the film has a slight pink tinge. For Treponema pallidum 

 stain from two to twelve hours. 



While the Romanowsky methods are more satisfactory for differential counts and 

 for the demonstration of the malarial parasites, and especially for differentiating 

 species, yet by reason of the liability to deterioration in the tropics of methylene 

 blue the hsematoxylin methods may be preferable. Many workers in blood-work 

 and cytodiagnosis prefer the haematoxylin. 



1. Fix the film either by heat with methyl alcohol for two minuetes or with Whit- 

 ney's fixative. Heat is to be preferred. 



2. Stain with Meyer's hemalum or Delafield's haematoxylin for from five to 

 fifteen minutes according to the stain. Frequently three minutes will be 

 found sufficient. To make the hemalum, dissolve 0.5 gram of hae matin in 25 

 c.c. of 95% alcohol. Next dissolve 25 grams of ammonia alum in 500 c.c. of 

 distilled water. Mix the two solutions and allow to ripen for a few days. The 

 stain should be satisfactory in two or three days. 



To make Delafield's haematoxylin, dissolve i gram of haematoxylin crystals in 

 6 c.c. of 95% alcohol. Add this to 100 c.c. of saturated aqueous solution of 

 ammonia alum. After exposure to light for a week, the color changes to a 

 deep blue-purple. Add to this ripened stain 25 c.c. of glycerine and 25 c.c. 

 of methyl-alcohol and, after it has stood for about two days, filter. The stain 

 should be filtered from time to time as a sediment forms. This makes a stock 

 solution which should be diluted 10 to 15 times with water when staining. 



Mink's Modification of TJnna's Haematoxylin. 



Haematoxylin, i gram. 



Alum, 8 grams. 



Sulphur (sublimed), i gram. 



Glycerine, 30 c. c 



Alcohol, 50 c.c.. 



Water, 100 c.c. 



Dissolve the hsematoxylin in the glycerine in a mortar. Dissolve the alum in the 

 water and add it to the glycerine haematoxylin in the mortar. Then add the sulphur 

 and the alcohol. The solution ripens in about three to four days. Allow the sedi- 

 ment to remain in the bottom of the bottle containing the stain and filter off small 

 quantities as needed. 



3. Wash for two to five minutes in tap water to develop the haematoxylin color. 



4. Stain either with a i to 1000 aqueous solution of eosin or with a 1/2 of i% 



