STUDIES ON CHROMOSOMES. 223 



In making the smear a bit of perfectly fresh, warm testis was 

 minced up into a fine pulp by means of a small-bladed scalpel 

 or with fine scissors and then spread into a thin film between two 

 slides in the same manner, that a blood film is prepared. The 

 slides after separation were plunged immediately into the fixing 

 agent where they remained from 30 minutes to several hours 

 depending upon the reagent used. At the end of this time they 

 were washed out in the appropriate liquid and all granules or 

 clumps of tissue which might prevent making a very thin, even 

 preparation were picked or scraped away. From this point the 

 slide was treated in the same way that ordinary sections are 

 treated. 



While many stains were tried none was found which surpassed 

 iron-haematoxylin for Gilson and Bouin material, or saffranin for 

 tissues fixed in Flemming or Hermann. The hcematoxylin 

 preparations were usually counterstained with orange G, acid 

 fuchsin or Congo red", although the- hsematoxylin alone was 

 found most satisfactory where photography was attempted. 

 Indeed some of my best preparations were found to be practically 

 worthless for photography because of a vivid red or yellow back- 

 ground which I was unable to eliminate by screens and which 

 therefore prevented adequate contrast in the photograph. In 

 preparations stained with safranin a counterstain of Lyon's 

 blue, lichtgriin, or Gentian violet was commonly employed. 

 I find a one per cent, safranin in anilin water a more satisfactory 

 stain than alcoholic solutions of safranin. 



While the haematoxylin preparations were far better than 

 safranin preparations for photographing, the latter often 

 revealed more detail in the mitotic figures because of the semi- 

 transparency of the chromosomes which often enabled one to 

 see separate elements where only a continuous black opaque mass 

 would be discernible with iron-haematoxylin. Delafield's haema- 

 toxylin gave satisfactory results with spireme and non-mitotic 

 stages but was of secondary value in the study of the fully formed 

 chromosomes. One set of smears stained first in safranin and 

 later in Delafield's haematoxylin proved to be unexpectedly 

 helpful in general study, although unfortunately such material 

 did not lend itself at all to photography. 



