HISTOLOGICAL EXAMINATION 19 



Eosin and methylene blue. The films must be fixed before 

 applying the staining solutions. Fixation may be secured by heat- 

 ing over the flame of a Bunsen burner or alcohol lamp, passing the 

 slide film side up through the flame until it is decidedly too hot 

 for the hand to bear then keeping it at this temperature for about 

 two minutes; or by heating in an oven at a heat of 110-120° C. for 

 5-10 minutes. Ether and absolute alcohol equal parts or absolute 

 alcohol alone is a satisfactory fixing agent. Ten to thirty minutes 

 are sufficient, though a somewhat longer time will not harm the 

 specimen. After the film is fixed it is covered with a saturated 

 alcoholic solution of eosin (Gruebler, ale. sol.) which is allowed to 

 act about 10 seconds. Then wash in water. The washing should 

 be rapid enough to give the red corpuscles a pinkish tint. With 

 slow washing they will be too deeply stained. As soon as the 

 eosin is washed off cover the film with a saturated aqueous solution 

 of methylene blue and let it act for one minute, then wash hastily 

 in distilled water. Shake off the water and dry quickly in the air. 



This stain is a good one though the fine granules of the poly- 

 morphonuclears are not stained. Jenner's, Wright's or Hasting's 

 stains are preferable. With eosin and methylene blue the red 

 corpuscles are pinkish, nuclei blue, eosinophile granules pink, 

 basophile granules deep blue. The fine granules of polymor- 

 phonuclears are unstained unless the specimen is much over stained 

 with eosin. The cell bodies of polymorphonuclears take a faint 

 pinkish tint. 



Giemsa's stain. The stain may be obtained already pre- 

 pared. It is made according to the following formula:— 



Azur II-eosin 3 gms. 



Azur II 0.8 gm. 



Glycerin (Merck C. P.) 250 gms. 



Methyl Alcohol (Kahlbaum 1) 250 gms. 



The blood film is dried in the air and fixed in absolute alcohol for 

 15 minutes or methyl alcohol for 2-3 minutes. Remove the al- 

 cohol with blotting paper, cover the preparation at once with the 

 freshly diluted staining fluid and let it act 10-15 minutes. The 

 staining fluid is diluted by adding one drop of the stain to 1 cc. 

 of distilled water in a small graduate and shaking gently. After 

 the diluted stain has acted 10-15 minutes wash the preparation 

 in a stream of distilled water. Remove the excess of water with 



