PRENTISS: THE OTOCYST OF DECAPOD CRUSTACEA. 179 
the fixative alone for three to five days, and washing out for at least two 
weeks in 90 % alcohol. The myelin sheath was intensely blackened, 
while all other tissues remained a yellowish brown. 
For tracing nerve fibres, both to peripheral and central endings, intra 
vitam staining proved of most value. Different methods were employed 
for obtaining peripheral and central stains. A one per cent solution of 
methylen blue in normal NaCl was injected into the body in either case. 
For peripheral endings several injections were made into the abdomi- 
nal blood space, at intervals of thirty minutes. When the animals 
showed signs of stupefaction, a final injection was introduced into 
the pericardial chamber. The amount of solution injected varied 
from a few drops, in Palemonetes, to five cubic centimetres, 
in the lobster. In from 15 to 30 minutes after the final injection 
the animals were usually dead. The part to be studied was then dis- 
sected out, barely covered with normal salt solution, and examined from 
time to time under the microscope, until a satisfactory degree of stain- 
ing had been reached. 
For central terminations one injection only was made, and this 
directly into the chamber of the heart, only a few drops of the solu- 
tion being required. When the blue color was well diffused throughout 
the tissues (about one hour after injection), the brain was dissected out, 
or exposed, and examined as before. For fixation of the stain Bethe’s 
ammonium-molybdate method for invertebrates was used. It was found 
to be better to leave preparations in xylol for only the shortest possible 
time, as this reagent diffuses the color. Preparations fixed by this 
method keep very well for a year or more, but after this they ultimately 
deteriorate, fibres originally sharp and continuous in outline becoming 
mere dotted lines, while the surrounding tissues take on a deep yellow 
hue. When both brain and otocyst were examined together, the peri- 
pheral cells and fibres stained first, then central fibres, central termina- 
tions, and ganglion cells of the brain in the order named. Sections 
60-120 uw in thickness were cut, but by far the greater number of pre- 
parations were examined in toto. The transparency of the tissues made 
this possible even with the brains, a millimetre or more in thickness, of 
large crayfish. 
To get constantly complete impregnations of both peripheral and cen- 
tral endings, it is necessary to expose to the atmosphere the part to 
be studied. The impregnation then takes place sooner, lasts longer, 
and affects a larger number of elements. The fixation of the color is 
also much better in this case, because the fluid can penetrate much more 
