CHROMOSOME NUMBER AND PAIRS IN AMBYSTOMA 175 
close to the spine. Under the binocular lens, in water, the 
peritoneum was carefully loosened from the underlying tissue 
by scraping it with a sharp scalpel, first along the edge cut 
from the back. Sections of this loosened edge were then grasped 
by the forceps and relatively large sheets were easily pulled off 
from the underlying tissue. The peritoneum covering the dorsal 
and lateral portion of the body wall is deeply pigmented and to 
it adhere considerable muscle and connective tissue when the 
peritoneum is removed. This portion was grasped with the 
forceps in removing the peritoneum from the body wall, as well 
as in all subsequent handling. Consequently the cells in the 
ventral transparent region available for study have been undis¬ 
turbed by instruments. However, there still remains a possi¬ 
bility that the strain of pulling the peritoneum loose might 
disturb some cells. 
Peritoneum fixed in Flemming’s and Hermann’s solutions was 
stripped from the body wall after four hours of fixation and then 
fixed twenty hours longer. That treated with the various picric 
acid mixtures was stripped immediately after fixation. However, 
the peritoneum fixed in the chromic acid modification of Bouin’s 
solution may be preserved in alcohol for as much as a year before 
stripping. That of Ambystoma punctatum, fixed in Flemming’s 
stronger solution and preserved in 5 per cent formalin, can be 
stripped at least six months after fixation. 
Material fixed in osmic acid fluids was washed five to fourteen 
hours in frequent changes of tap-water. Picric acid preparations 
were gradually transferred to 70 per cent alcohol, beginning with 
10 per cent and progressing through successively stronger grades 
differing by 10 per cent. They remained in each grade five to 
ten minutes. The tissues remained in 70 per cent alcohol con¬ 
taining a few drops of saturated aqueous lithium carbonate 
solution until the picric stain was removed, and before staining 
they were returned to water by reversing the above process. 
All of the material was stained in Heidenhain’s haematoxylin after 
mordanting in a per cent solution of iron alum for four to six 
hours. No counterstains were used. 
JOURNAL OP MORPHOLOGY, VOL. 33, NO. 1 
