BIGELOW: EARLY DEVELOPMENT OF LEPAS. 67 
Eggs prepared by the above method were so transparent that even in 
later stages the outlines of cells on either side of the embryo could be 
clearly seen by appropriate focussing. It was, therefore, easy to study 
and draw optical sections in any plane. The refractive index of clove 
oil! is such that the vitelline membrane becomes almost invisible. 
By carefully moving the cover glass it is possible to roll eggs into any 
desired position, and for this purpose the balsam was for months kept 
semi-fluid by occasionally applying a drop of xylol to the edge of the 
cover glass. 
It was found practicable, and in some cases profitable, after studying 
an egg in balsam, to remove the cover glass, dissolve the surrounding 
balsam with xylol, lift the egg by means of a capillary tube, transfer it 
to paraffine, imbed by the watch-glass method and section it. When 
imbedded near the surface of the block of paraffine, the long axis of the 
egg can be distinguished by the use of a lens, and hence sections can be 
cut longitudinally or transversely as desired. This method of sectioning 
single eggs was employed only for the purpose of gaining an idea of the 
appearance of sections of particular stages in known planes. Asa rule, 
pieces of the egg-lamellz rather than single eggs were imbedded and 
sectioned, the sections being stained on the slide. Since the eggs have 
no definite arrangement in the lamelle, sections in all planes were thus 
obtained. By comparison with sections of single ova in which the orien- 
tation had been definitely established, it was possible to choose with 
certainty the sections representing any desired plane in any stage of 
development. 
For staining sections on the slide Delafield’s hematoxylin diluted 
with four or five times its volume of distilled water gave the best results. 
In the later cleavage stages and in embryonic stages orange G or eosin 
were used after the hematoxylin. By this means the entoblastic yolk- 
cells were sharply differentiated. 
T» the study of preparations of the entire eggs a sub-stage condenser 
with iris diaphragm was absolutely necessary. A 7, inch homogeneous 
immersion objective with long working distance was of great service. 
Most of the preparations upon which this paper is based are yet in 
good condition, and are therefore available as evidence in support of the 
following account of the development of Lepas. 
1 Since this paper was written I have found that oil of cassia for clearing gives 
results even superior to those obtained by the use of clove oil. It has also proved 
to be an excellent mounting medium, but probably the preparations will not retain 
stains permanently. 
