No. 2.] THE OVARIAN EGG OF LIMULUS. I 15 



Imbedding has been done in the usual way by means of par- 

 affine. The mature eggs of the ovarian tubes were imbedded 

 and sectioned in celloidin. To enable the imbedding medium 

 to penetrate, a slit was made in the chorion by means of a 

 sharp razor. 



Previous to imbedding, the absolute alcohol was removed by 

 means of chloroform, saturated with dissolved paraffine. To 

 avoid the hardening effect of the chloroform on the yolk of 

 the larger eggs, xylol and turpentine were substituted for the 

 chloroform. 



The paraffine sections, from five to ten \x, in thickness, were 

 fixed to the slide by means of water, Mayer's albumen fixative, 

 or by the two combined. 



Staining was done almost exclusively after the sections were 

 mounted on the slide. The sections of the larger eggs in cel- 

 loidin were stained with Delafield's haematoxylin diluted ten 

 times with water and slightly acidulated with HCl. This 

 leaves the yolk spheres unstained and facilitates the search 

 for traces of the nucleus and the maturation spindle. 



For the study of karyokinesis, Heidenhain's iron-haematoxy- 

 lin was used. The archoplasm and centrosome in the younger 

 eggs were studied by means of Heidenhain's iron-haematox- 

 ylin, either alone or followed with erythrosin, eosin, or acid 

 fuchsin. Erythrosin and cyanin have also been used to good 

 advantage ; also borax-carmine, followed with picric acid ; Del- 

 afield's haematoxylin, either alone or followed with picric acid ; 

 Weigert's picro-carmine ; Ehrlich's haematoxylin, either alone 

 or followed with erythrosin, eosin, and especially with acid 

 fuchsin ; eosin and nigrosin to a limited extent ; the Biondi- 

 Ehrlich mixture, and finally lithium-carmine and Lyon's blue. 



These stains all give good results, but they differ in the 

 extent to which they can be applied. The carmine stains 

 have not been found useful on material killed in Merkel's or 

 Flemming's fluids. To obtain the double stain with Lyon's 

 blue, safranin has been substituted for the carmine on Mer- 

 kel's material. In such cases the sections were first stained in 

 Lyon's blue for twenty-four hours, after which they were 

 stained for twenty-four hours in safranin ; and previous to 



