280 Dr. A. B. Macallum. On the Demonstration 



alcoholic solution of ammonium sulphide for twenty-four hours, and 

 kept at the temperature of the laboratory. The pieces of tissue 

 remained for two weeks or more in the warm ammonium sulphide, 

 but no colour reactions appeared in the chromatin of the cells. Not 

 satisfied with the conditions under which the experiments were 

 carried on, I adopted other methods. It occurred to me that, if I kept 

 a few cells for a long time completely separated from one another under 

 a cover-glass, on a slide and surrounded with ammonium sulphide, 

 the reaction might come out. To prevent the evaporation of the 

 ammonium sulphide, I luted the edges of the cover to the slide with 

 various luting compounds, only to find that, in some way or other, 

 the preparation spoiled after a day or two. The luting method being 

 useless, I employed another device. Glycerine, when kept in contact 

 with ammonium sulphide at a moderately high temperature for some 

 time, does not affect the latter reagent, and is itself unaffected. I 

 teased out with clean goose-quill points on a slide a small piece of the 

 testicle of Necturus, hardened in 70 per cent, alcohol, added a drop 

 of freshly-prepared ammonium sulphide, put on a cover-glass, allowed 

 a drop of glycerine to run in from its edge, and then placed the slide 

 in a warm oven with a constant temperature of 60 C. Here it was 

 allowed to lie for three days, at the end of which time I examined it 

 under the microscope, and found that, in addition to that reaction 

 distinctly present in the cell body and nucleus of each red corpuscle, 

 there was one apparent in the nuclei of a group of testicular cells at 

 one spot in the preparation. The slide was replaced in the warm 

 oven, and the daily examination of it showed that, accompanying the 

 increase in the number of cells presenting the iron reaction, there 

 was an increase in the depth of the colour in those nuclei first 

 affected, until, at the end of twenty days, the great majority of the 

 testicular nuclei under a cover-glass, 16 mm. square, manifested a 

 colour varying from light green or greenish-blue to dark-green or 

 black. Under a high-power objective the colour was found confined 

 to the chromatin nodules and nuclear network. After three weeks 

 the nuclei adjacent to the edges of the cover-glass began to lose their 

 stained appearance, until, finally, the chromatin possessed only a rusty 

 appearance due to the formation of ferric oxide, for, when a mixture of 

 hydrochloric acid and potassic ferrocyanide was allowed to run under 

 the cover, the rust-coloured nuclei immediately assumed a deep azure- 

 blue colour. That it was the chromatin alone in such cells which 

 presented the reaction with ammonium sulphide was abundantly 

 shown in the karyokinetic figures present in the same preparation. 

 The achromatin and cell substance were unaffected. 



Encouraged by the success of this experiment, I made a number of 

 preparations from the other organs of Necturus, hardened also in 70 

 per cent, alcohol. Nearly all of these were successful, but the time 



