107 



another test'- tube. The filtrate should be perfectly clear, 

 not cloudy. To the anilin water thus obtained a satu- 

 rated alcoholic solution of gentian violet is added till the 

 iluid is deeply colored. 



Some of the anilin water-gentian violet thus pre- 

 pared is poured out into a watch-glass. A streak cover - 

 glass preparation of anthrax is now carefully fixed in tlie 

 flame. Care must be taken not t.o over-heat the specimen, 

 as the anthrax bacillus when over-heated does not stain 

 satisfactorily. The fixed cover-glass is then placed 

 between the thumb and forefinger, with the specimen side 

 down, and carefully dropped upon the surface of the stain 

 in the watch-glass. It is then allowed to float on the dye 

 for 10 to 15 minutes. Sometimes it is necessary to warm 

 the dye on the radiator or on an iron plate in order to 

 obtain a rapid and intense stain. The cover-glass is then 

 picked up with the forceps, thoroughly washed with 

 water, and immersed in a solution of iodine in potassium 

 iodide. This is made by dissolving 2 g. of potassium 

 iodide and 1 g. of iodine in 300 c. c. of distilled water. 

 The specimen is allowed to remain in the iodine for ^ to 

 1 minute, or even several minutes. Care must be taken 

 not to expose too long to the action of iodine, as it tends 

 to contract the protoplasm into granules. The cover-glass 

 is then removed from the iodine, washed with water, and 

 moved about in a watch glass of strong alcohol, to which, 

 if necessary, a drop of acetic acid may be added, to assist 

 the decoloration. From time to time the cover-glass 

 should be washed with water and examined with No. 7 

 objective to ascertain the progress in decoloration. When 

 finally a colorless back-ground is obtained for the deeply 

 stained violet bacilli the washing in alcohol is discontin- 

 ued. The cover-glass is then washed with water and 

 stained with dilute eosine for ^ to ^ minute. The eosine 

 is an acid anilin dye, and therefore stains the protoplasm 

 of cells, nuclei, etc., but not bacteria. Care must be taken 

 not to overstain the preparation with eosine, as it would 



