i70 HOPE HIBBARD 



water. The eggs of this form are mature when shed or when they 

 easily shake out of the ovary, and may be fertihzed by species 

 sperm as soon as they are clean, but eggs need further treatment 

 before cross-fertilization. To clean from bits of ovarian tissue 

 and coelomic fluid, the water in the finger-bowl in which the 

 ovaries have been shaken is stirred, then allowed to settle for a 

 brief period and the supernatant water poured off. The finger- 

 bowl is then refilled with fresh sea-water and the process re- 

 peated several times. In this way the heavier eggs which settle 

 are kept, while the hghter debris is poured off. Since these 

 eggs were not normally fertilizable by Arbacia sperm, it was 

 necessary to resort to some artificial means of breaking down the 

 cortical resistance. As in other eggs, there are three methods 

 of breaking down this resistance, namely, 1) by stahng the eggs, 

 2) by over-insemination, and, 3) by the use of alkah. For 

 the purpose of this work the third method was employed. To 

 determine the optimum concentration of alkah, two experiments 

 were performed. Varying amounts of n/10 NaOH in one case 

 were added to sea-water, and n/10 KOH in the other. After 

 fertihzation by Arbacia sperm, the percentage of development 

 was recorded. Very little difference was found between the 

 NaOH and the KOH. The standard strength of twenty drops 

 of n/10 NaOH per 150 cc. of sea- water was adopted as the pro- 

 portion of alkali yielding the best results. The alkaUne sea- 

 water was added to the cleaned eggs from which most of the water 

 had been poured, and as soon as possible the sperm suspension 

 wss added and the contents of the dish well mixed. As soon as 

 the eggs had settled, the supernatant hquid containing excess 

 sperm and alkah was drawn off by means of a suction flask con- 

 nected as figured. 



By this method all the hquid save about 8 cc. could be quickly 

 withdrawn, leaving the eggs undisturbed on the bottom of the 

 finger-bowl. Fresh sea-water was then added. After self-fer- 

 tihzation the supernatant water was similarly withdrawn and 

 replaced by fresh sea-water. 



In any one series of eggs preserved the eggs from only one 

 female were used and ihey were fertihzed by sperm from one 



