MICROSCOPIC EXAMINATION 131 



gol's solution (iodine 1, potassium iodide 2, water 100) for 

 one minute, rinse in water, blot to remove excess of water, 

 dehydrate in aniline oil (or better aniline oil 2 parts and 

 xylene 1 part) clear in xylol and mount in balsam. 



Staining spores. Make a cover-glass preparation, dry, 

 and flame as already described. Take the preparation by the 

 edge with the fine forceps, * cover the film surface with carbol 

 fuchsin, and hold the preparation over the gas flame until 

 steam is given off ; then remove it for a few seconds and heat 

 again. Repeat the heating three or four times. After the 

 stain has acted for from 3 to 5 minutes, rinse the preparation 

 in water and decolorize it by immersing it in a watch glass 

 containing about 3 cc. of 1% solution of sulphuric acid or 

 95% alcohol. After about one-half minute remove the prepa- 

 ration and rinse it thoroughly in water. If it is not decolor- 

 ized, repeat the bleaching process. This removes the coloring 

 matter from the bodies of the bacteria, but leaves it in the 

 spores. After thoroughly washing the preparation, counter- 

 stain it with a saturated aqueous solution of methylene blue 

 for about 30 seconds, rinse in water, and examine. The spores 

 should be stained red (with the fuchsin) and the rest of the 

 organism should be colored blue. 



Moeller's method is designed still further to favor the 

 penetration of the coloring matter through the spore mem- 

 brane. The prepared cover-slip is held for two minutes in 

 chloroform, then washed off in water, then placed from one- 

 half to three minutes in a 5 per cent solution of chromic acid, 

 again washed in water, and stained in dilute (1 to 10) carbol 

 fuchsin, heated slowly over flame until steam is given off, for 

 3 to 5 minutes. The staining fluid is then washed off and the 

 preparation decolorized in a 3 per cent solution of hydro- 

 chloric acid or a 5 per cent solution of sulphuric acid. Wash 

 and stain for a minute in aqueous methylene blue. The spores 

 will be red and the body of the bacteria blue. Different spores 

 vary greatly in the readiness with which they take up the 

 dyes, and we have, therefore, to experiment with each species 



