ArmFIClAL PAUTIIFNOGICNFSIS IN ECHINUS ES(!ULENTUS. 543 
is exceedingly higli. Tliere is no membrane formation by 
Delage’s method, and tliis, together with the characters of 
the larvfB mentioned above, forms a parallel to the I'esults 
obtained by Loeb ( 11 ) when using hypertonic sea-water 
alone as a method of starting segmentation. 
V. Combined Method. 
This suggested to ns that possibly if the eggs of E. 
escnlentus were treatetl, first with butyric acid to bring 
about membrane formation, and then with tannic acid and 
ammonia, we might succeed in rearing large numbers of 
healthy larvae. This method, which is a combination of those 
elaborated by Loeb and Delage, has been the most successful 
at Plymouth this season (1912). Table XIII shows the 
relative proportions of blastulie obtained by it, as compared 
with the numbers obtained by either of the other two 
methods. 
4’able XIII. 
(1) 3 c.c. N/10 hiityric acid -|- flO c.c. sea- water 
for 1'5 min. 
10 c.c. sea- water -)- 40 c.c. sugar 10 c.c. 
M/OO tannic acid for 0 min. 
I'l c.c. N/10 ammonia added for 1 hour 
(2) 3 c.c. N/10 Inityric acid -(- 50 c.c. sea-water 
for 1'5 min. 
lO c.c. sea-water -|- 40 c.c. sugar -f- 2‘0 c.c. 
M/OO tannic acid for 0 min. 
21 c.c. N/10 ammonia added for 1 hour. 
(3) 10 c.c. sea-water + 40 c.c. sugar -f- 1'4 c.c. 
M/00 tannic acid for 0 min. 
1'5 c.c. N/10 ammonia for 1 hour. 
(4) 3 c.c. N/10 butyric acid + 50 c.c. sea- water 
for 1'5 min. 
•2 c.c. N/10 NaOH -|- 50 c.c. sea- water for 
0 min. 
8 c.c. 2‘5 M NaCl -|- 50 c.c. sea-water for 
45 min. 
80 per ceiit. blastula; 
swimming vigor- 
ously and very 
healthy in aj^pear- 
ance. 
75 per cent, blastuhc. 
70 per cent, blastuhc. 
00 per cent, blastulac 
