CASTLE: EMBRYOLOGY OF CIONA INTESTINALIS. 215 
them to alcohol of a low grade, or to water, the egg envelopes would 
again stand out clear of the ovum, as in the living egg. By then suck- 
ing the eggs one at a time into a glass tube of the proper calibre, the 
entire envelope, consisting of follicle cells, test cells, and chorion, could 
be removed. with considerable facility, and in the majority of cases with- 
out injury to the egg itself. Eggs thus decorticated and then mounted 
afforded excellent surface views. 
The eggs are rather opaque, on account of the large amount of yolk 
which they contain, so that any stain except a very faint one is an ob- 
stacle in the study of whole preparations. Excellent results were obtained 
by mounting in balsam, without any staining whatever, eggs which had 
been killed in Perenyi's fluid and decorticated.! But for the 64-cell and 
later stages staining was found desirable. Many carmine and hematoxy- 
lin stains were tried ; the one which gave by far the best results being 
Orth's picro-carminate of lithium. The eggs were treated with a small 
amount of this stain in a watch-glass for from six to twenty-four hours, 
then washed thoroughly in water. By this method resting nuolei are 
Stained bright rose-color, while all other structures take only a faint 
yellow color from the picric acid, and give up even this if the washing is 
sufficiently prolonged. But the carmine stain in the nuclei is extremely 
tenacious, and does not fade in the least upon prolonged washing in water 
or preservation for months in strong alcohol. After the eggs had been 
stained and decorticated, they were dehydrated, cleared in xylol or cedar 
oil, then mounted in balsam, the cover glass being supported with small 
glass rollers made from fine capillary tubes. These served the double pur- 
pose of preventing the orushing of the egg and allowing it to be rolled 
into any desired position by movement of the cover glass. Changing the 
position of the egg, however, is not often necessary, for at an early stage 
it takes on a flattened form, which causes it to come to rest with the dorsal 
or the ventral surface uppermost. This is the case at all periods between 
the 24-cell stage and that at which the neural tube begins to elose, ex- 
cept for a brief period, when the embryo consists of from forty-six to 
sixty-four cells, and the vertical axis becomes equal to or even greater 
than the longitudinal axis. Then there is no single position of stable 
repose for the embryo, and rolling is often necessary to bring it into the 
positions desired. 
The self-orientation of the egg during most of the early stages was of 
1 T find that Lillie (95) has obtained good results in the case of the eggs of the 
mollu.k Unio by mounting, without staining, material killed in Perenyi’s fluid. 
He, how^ver, used glycerine instead of balsam as a mounting medium. 
