98 BIOLOGY AND TECHNIQUE 



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

 in the flame. 



Wash in chloroform for two minutes. 



Wash in water. 



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



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

 tion 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. 2 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. 3 (1) Copper Sulphate Method. Cover-slip prepara- 

 tions are made by smearing the organisms in a drop of animal serum, 

 preferably beef-blood serum. 



Dry in air and fix by heat. 



Stain for a few seconds with 



Saturated alcoholic solution of fuchsin or gentian-violet 5 c.c., in 

 distilled water 95 c.c. 



The cover-slip is flooded with the dye and the preparation held for a 

 second over a free flame until it steams. 



Wash off dye with twenty per cent aqueous copper sulphate solution. 



Blot (do not wash). 



Dry and mount. 



By this method permanent preparations are obtained, the capsule 

 appearing as a faint blue halo around a dark purple cell body. 



(2) Potassium Carbonate Method. This method consists in using 

 as a dye a half -saturated solution of gentian- violet. Gentian-violet in 



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



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



' Hiss, Cent. f. Bakt., xxxi, 1902; Jour. Exper. Med., vi, 1905. 



