STUDIES TN SPICULE FORMATION. 51 



Naples and preserved in alcoliol. In all cases I embedded 

 portions of the colony in paraffin wax and cut thin free- 

 hand sections with a razor ; I then stained these sections in 

 the manner described below. The spicules are identical in 

 the three species. Staining methods which do not differ- 

 entiate the nuclei very prominently do not give good results 

 for ascertaining the number of scleroblasts in connection 

 with the adult stellate spicules, chiefly because the mass of 

 the spicule more or less effectually hides all objects situated 

 underneath, and the conical protuberances largely obscure 

 objects situated to the side. The use of picro-carmine would 

 doubtless give better results, but I was unable to employ this 

 stain. The only effectual method which I employed was the 

 ordinary borax-carmine method, the differentiation with acid- 

 alcohol dissolving the spicules sufficiently to render the 

 scleroplasm apparent. This method merely revealed one 

 scleroblast (nucleus) in connection with each spicule, the 

 layer of scleroplasm closely investing the entire spi- 

 cule and following all its outlines, and the large nucleus 

 being situated in a mound of protoplasm at the periphery 

 (figs. 17, 18). The stellate spicule originates in a scleroblast 

 as a spherical granule (fig. 14 ; these early stages are quite 

 visible in the non-decalcified safranin and lichtgriin-stained 

 preparations) which later acquires spines on its surface as 

 it increases in size (fig. 16), and which spines ultimately 

 become the conical processes of the adult spicule. 



Previous literature dealing with the scleroblastic develop- 

 ment of Ascidian spicules is very small in amount (see 

 Herdman [2] for the literature relating to Ascidian spicules 

 up to 1885, since which year, I believe, no literature on the 

 present subject has appeared). Loewig and Kolliker [6] in 

 1846 provided very poor figures of the stellate " cellules 

 incrustees" in Didemnum, and illustrated one of these 

 " cellules " partly decalcified, showing the cell-wall spherical 

 in outline. Giard [1] in 1872 figured stellate spicules of 

 Euccelium and Didemnum also situated each within a circle 

 which is supposed to represent the cell-wall; he figured as 



