238 THE CYCLE OF CELL-DIVISION 



chloroform. It was found that the constituent which 

 induces divisions in lymphocytes is almost entirely 

 confined to the azur dye. The more the azur was 

 extracted the less efficient the polychrome dye became, 

 and the azur is very potent although it does not stain 

 the chromatin as well as the polychrome dye. 



A concentrated solution of azur dye was made 

 thus: In a burette 20 cc. of Unna's polychrome methy- 

 lene blue (Grubler) had added to it 20 cc. of chloroform, 

 and the mixture was allowed to stand for 12 hours. 

 The chloroform, which sinks to the bottom, carrying 

 some of the azur dye with it, was then run off into 

 a shallow dish, where it was allowed to evaporate. 

 20 cc. more of chloroform was then added to the 

 original 20 cc. of stain in the burette, and, after 

 12 hours, it, in its turn, was run off into the same 

 dish and also allowed to evaporate. This procedure 

 was repeated five times, and the dry azur dye was 

 so obtained. Lastly, a solution of this dye was made 

 by adding 5 cc. of water to the dish. This potent 

 dye is a fluorescent red one, which, when dry, shines 

 with a metallic lustre. A very powerful jelly for 

 causing mitotic divisions in lymphocytes was made by 

 substituting . 4 cc. of this potent solution for the . 2 cc. 

 of Unna's stain in the last equation. By means of this 

 <jelly all stages of divisions can be readily obtained, it 

 being only necessary to vary slightly the content of 

 alkali in producing early or late mitosis in the ten 

 minutes. It is better to keep at least two units of 

 polychrome stain in the jelly, in order to stain the 

 chromosomes more deeply. 



