J. R. SHAVER 267 



fuged eggs was made by Harvey by means of staining with methyl 

 green which she considers "just as specific" as Janus green. Shaver 

 ( unpubhshed observations ) centrifuged Lytechinus pictus unfer- 

 tihzed eggs into "hght" and "heavy" halves, and the former into 

 "clear" and "granular" quarters, with the same force of gravity 

 employed by Harvey. These fragments were observed with phase 

 contrast microscope, unstained, or after staining with Janus green 

 or methyl green. The fine line of granules seen in the clear quar- 

 ters under oil immersion dark phase objective appears to be a 

 mixture of elements. The largest were spheres or spheroids of 

 from 0.5 to 1.0 micron diameter. Harvey (1946) gives the dimen- 

 sions of mitochondria seen in granular quarters and heavy halves 

 as 0.6-1.0 micron. In some of these particles observed by Shaver, 

 the bipartite structure seen frequently in mitochondria of sea ur- 

 chin embryonic cells, as well as in adult tissue cells of mammals 

 (Harman, 1950), was unmistakable. The other elements of the 

 fine line of granules (just at the limit of visibility at 1400 X mag- 

 nification) consisted of very much smaller granules and very 

 small rods. The latter appeared to be closer to the oil cap, and in 

 some fragments formed a boundary between it and the granular 

 stratum just below. 



The results of staining with Janus green or methyl green were 

 not very definitive. The former stain has not proved satisfactory 

 for mitochondria in the intact cells of sea urchin embryos ( Shaver, 

 1955). In fragments of centrifuged eggs, granular elements cor- 

 responding to mitochondria ( in the sense used by Harvey, 1946 ) , 

 as well as the elements of the "fine" layer of granules in the clear 

 quarter, and a portion of the yolk layer, all took on a bluish-green 

 to purple color in the presence of the dye; phase contrast observa- 

 tion of stained fragments also indicated that there was a physical 

 change in the granular elements after dye penetration, a tendency 

 to clumping being seen, especially in the granular quarters. 

 Methyl green proved unsatisfactory as a specific stain for mito- 

 chondria since, under the conditions in which it was used, most 

 of the granular elements in all quarters, or halves, stained a bluish 

 purple. 



The clear quarters were unfortunately not followed after ferti- 



