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 1 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 paramne, imbed by the watch-glass method and section it. When 

 imbedded near the surface of the block of paramne, 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. As a rule, 

 pieces of the egg-lamellse rather than single eggs were imbedded and 

 sectioned, the sections being stained on the slide. Since the eggs have 

 no definite arrangement in the lamellae, 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 haematoxylin 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 haematoxylin. By this means the entoblastic yolk- 

 cells were sharply differentiated. 



In the study of preparations of the entire eggs a sub-stage condenser 

 with iris diaphragm was absolutely necessary. A -^ 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. 



