Life-History of Isoetes. 233 
After the specimens were imbedded they were sectioned 
with a Minot-microtome, stained on the slide, and mounted in 
Canada-balsam. While safranine and gentian-violet, in water, 
give a good nuclear stain, the young cell-walls colour but 
little, and often do not show as clearly as could be wished, 
Bismarck-brown, however, particularly when dissolved in 70 
per cent, alcohol, stains the young cell-walls especially well, 
and is by far the best stain for this purpose that I have yet 
found. Extremely beautiful preparations of the older pro- 
thallia and embryos were made by staining in toto with 
alum-cochineal or Kleinenberg’s haematoxylin, for a nuclear 
stain, and then staining on the slide with the Bismarck-brown. 
In the younger stages, however, nearly all attempts at staining 
in toto were ineffectual. Bismarck-brown is especially satis- 
factory also, in that it does not overstain, and is not readily 
extracted by alcohol, so that there is not the danger of too 
much decoloration in dehydrating the specimens that is met 
with when using other aniline colours. 
I am especially indebted to Mr. F. V. Coville of Washington, 
through whose kindness a very abundant supply of fruiting 
specimens of Isoetes echinospora , var. Braunii , Durieu, was sent 
me. The plants were collected in the vicinity of Washington 
in September, 1889, and at this time the spores were ripe, or 
nearly so. The plants were kept in an aquarium with about 
an inch of sand at the bottom, and remained in good condition 
through the winter, resuming growth vigorously the following 
spring. The spores are ordinarily set free by the decay of the 
sporangium-wall, and this does not occur, probably, until late 
in the autumn or winter, when the plants are growing under 
normal conditions ; so that probably the spores do not ger- 
minate naturally until spring. If, however, they are set free 
artificially, they will begin to germinate very soon, but the 
process is slower than when set free spontaneously. 
The first sowings were made by me Sept. 27, and from 
that time on, at intervals, through the autumn and winter. 
There was much difference shown in the promptness of ger- 
mination, even in the same lot ; and as the absolute opacity 
