EXAMINATION OF BACTERIA WITH THE MICROSCOPE. 47 



cent, aqueous solution of water soluble yellow esoin. To this 

 mixture add 70 c.c. or 80 c.c. of a i per cent, aqueous solu- 

 tion of methylene-blue (Ehrlich rectified). Filter. Dry the 

 residue left on the filter paper, powder in a mortar. About 

 .7 to i gram of powder is thus obtained. Dissolve in about 

 400 c.c. of pure methyl alcohol (Merk). 



Nocht* recommends the addition of silver oxide to i per 

 cent, methylene-blue solution. The silver oxide in this case 

 is obtained by precipitating i gram of silver nitrate with suf- 

 ficient alkali, and adding this to 100 c.c. of the i per cent, 

 methylene-blue solution, and allowing the mixture to ripen for 

 four or five days at room temperature. 



Goldhornf recommends the use of lithium carbonate in- 

 stead of the sodium salt. His directions are to boil 2 grams of 

 lithium carbonate, and 2 grams of methylene-blue in 200 c.c. of 

 distilled water. This is done in a double boiler, and from time 

 to time samples are examined in test-tubes to see whether the 

 polychrome is formed as indicated by a red color. The mix- 

 ture after cooling spontaneously is filtered, and the half of it 

 is rendered distinctly acid with acetic acid. The two halves 

 are then poured together. The rest of the procedure does not 

 differ essentially from others described except that he recom- 

 mends commercial wood alcohol in the place of pure methyl 

 alcohol for dissolving the stain. 



Wright's modification is as follows: To 100 c.c. of a i per cent, solution of 

 sodium bicarbonate in water add i gram of methylene-blue. Place in the 

 steam sterilizer at 100 C. for one hour. When cool add one-tenth per cent, 

 watery solution of eosin (Griibler, yellowish, soluble in water) until the mixture 

 loses its blue color, becomes purple, and a metallic scum forms on the surface. 

 About 500 c.c. of the eosin solution are needed. Collect the precipitate on a 

 filter; let it dry; make a saturated solution of the precipitate in methyl-alcohol; 

 filter. To the quantity obtained add one-fourth as much methyl alcohol, so 



*Enzyklopaedie d. Mikros. Technik., quoted by Hastings, Jour. Exper. 

 Med.,Vol. 7, 1905, p. 266. See also Duval. Jour. Exper. Med., IX., 1907, 

 P- 381- 



\Jour. Exper. Med., Vol. VIII., 1907, pp. 451-459. 



