150 MICROMETRY AND BLOOD PREPARATIONS 



In Leishman's method the polychroming is accomplished by adding 

 i gm. of methylene blue to 100 c.c. of a 1/2% solution of sodium car- 

 bonate. This is kept at 65 C. for 12 hours and allowed to stand at 

 room temperature for 10 days before the eosin solution is added. The 

 succeeding steps are as for Wright's stain. 



The modification of the Romanowsky stain, which is used in the 

 laboratory of the U. S. Naval Medical School and which can be rec- 

 ommended as giving good results with the least expenditure of time 

 in making and which by the addition of either acid or alkaline alcohol 

 can be made to give the staining effect desired, is that of Hospital 

 Steward R. W. King, U. S. Navy. The preparation of the stain is as 

 follows : 



Dissolve i gram of Grubler's methylene blue and 0.5 gram of 

 sodium, bicarbonate in 100 c.c. of distilled water. Transfer to porce- 

 lain dish and evaportate to dryness over Bunsen burner or alcohol 

 flame. The fluid may be allowed to boil gently until about half of the 

 water has escaped, when the heat should be somewhat reduced. 

 Evaporation is now facilitated by causing the fluid to flow upon the 

 sides of the dish by tilting. This also overcomes the tendency to 

 spluttering. The heat should be continued until the last trace of 

 moisture has disappeared. The absolutely dry stain is then removed 

 from the dish and preserved in small well-stoppered vials. 



(If larger quantities than the above are to be made, separate dishes 

 must be used.) 



Stock solution: Dissolve 0.3 gram of the polychromatic blue and 

 0.175 gram of Grubler's eosin in 100 c.c. of pure methyl alcohol. 

 Allow to stand for three hours and filter. Add 25 drops of 3% hydro- 

 chloric acid alcohol (used in T. B. staining). 



For use, take 25 c.c. of the stock solution and add eight drops of the 

 alkaline alcohol (5 c.c. 10% sol. caustic soda to 100 c.c. 95% alcohol), 

 and test by staining a section of a fresh blood film. If the blue over- 

 stains, add one drop of the acid alcohol and test again. If the eosin 

 overstains (nuclei stain poorly), add two or three drops of the alkaline 

 alcohol. In this way it will only require one or two trials to adjust 

 the staining properties of the fluid, after which it will keep unchanged 

 for some time. At any time, however, if it is found not to give good 



