78 JouRNAL OF CoMPARATIVE NEUROLOGY. 
b. Intra-Vitam Injection.—The coloration of the nerve- 
cells through intra-vitam injection of methylen-blue was given 
a most thorough trial, a large number of animals being utilized 
for this purpose. The subcutaneous injection preferred by 
MEYER (96) is not practicable for Mustelus because of the 
absence of either loose areolar tissue or of lymph spaces. The 
syringe was therefore inserted directly into the vascular system. 
A 5% solution of methylen-blue, BX brand, was injected some — 
four times during the course of an hour. Beginning witha 
small quantity, the amount rose successively until as much as 
30 c.c. was introduced in the final injection, making some 
50 c.c. in all. This whole process was governed, however, 
not by fixed quantities of the reagent nor by exact periods of 
time, but by the stopping of the heart’s action and the blueness 
of the animal. Half an hour after the final injection, the brain 
was removed, cut into thin slices, and then exposed to the air 
until the tint had become a brighter blue. In the conversion 
of the unstable methylen-blue stain of the fresh tissues into the 
insoluble form, I have not been successful with the method 
recommended by BrTuE (’96). The use of the picrate of am- 
monia as a preliminary fixer has seemed to actually impair the 
clearness of the final preparation. I obtained the best results 
with the solution given by MEYER (’96): 
Distilled water ‘ : : 100 C.c. 
Ammonium molybdate. | IO grams 
Hydrochloric acid . : : 10 drops 
Heat the first two ingredients together, then add the acid. 
The pieces of brain were placed in this mixture, cooled 
with ice, for four hours. They were then washed with iced 
water for two hours. Dehydration with cooled alcohols, and 
imbedding in paraffin were hastened as much as practicable ; 
in fact, it is well to have the tissues in paraffin on the same day 
when the injection was begun. The preparations were used 
chiefly for the study of the architectural relations between the 
neurones, and so the sections were cut quite thick. 
The results given by this method are characterized by ex- 
ceptional clearness, due, in large measure, to the selective col- 
