116 BIOLOGY AND TECHNIQUE 



SPECIAL STAINING METHODS 



Spore Stains. ABBOTT'S METHOD. 6 Cover-slips are smeared and fixed by 

 heat in the usual manner. 



Cover with Loeffler's alkaline methylene-blue and heat the stain until it 

 boils, repeat the heating at intervals but do not boil continuously. Keep 

 this up for one minute. 



Rinse in water. 



Decolorize with a mixture of alcohol eighty per cent 98 c.c. and nitric 

 acid 2 c.c., until all blue has disappeared. 



Rinse in water. 



Dip from three to five seconds in saturated alcoholic solution of eosin 

 10 c.c., and water 90 c.c. 



Rinse in water, blot, and mount in balsam. 



By this method the spores are stained blue, the bodies of the bacilli are 

 stained pink. 



MOELLER'S METHOD/ Cover-slips are prepared as usual and fixed in the 

 flame. 



Wash in choloroform for two minutes. 



Wash in water. 



Coyer with five per cent chromic acid one-half to two minutes. 



Wash in water. Invert and float cover-slip on carbol-fuchsin solution in 

 a small porcelain dish and heat gently with a flame until it steams; continue 

 this for three to five minutes. (This step can also be done by covering the 

 cover-glass with carbol-fuchsin and holding over flame.) 



Decolorize with five per cent sulphuric acid five to ten seconds. 



Wash in water. 



Stain with aqueous methylene-blue one-half to one minute. By this 

 method spores will be stained red, the body blue. 



Capsule Stains. WELCH'S METHOD. S Cover-slips are prepared as usual 

 but dried without heat. 



Cover with glacial acetic acid for a few seconds. Pour off acetic acid 

 and cover with anilin water gentian-violet, renewing stain repeatedly until 

 all acid is removed. This is done by pouring the stain on and off three or 

 four times and then finally leaving it on for about three minutes. 

 Wash in two per cent salt solution and examine in this solution. 

 Hiss' METHODS. 9 (1) Copper Sulphate Method. Cover-slip preparations 



e Abbott, "Prin. of Bact.," Phila., 1905. 



7 Moeller, Cent, f . Bakt., I, x, 1891. 



8 Welch, Johns Hopkins Hosp. Bull., 1892, iii, 81. 



8 Hiss. Cent. f. Bakt., xxxi, 1902; Jour. Exper. Mod., vi, 1905. 



