202 ARTIFICIAL PARTHENOGENESIS AND FERTILIZATION 



an hour and frequently shaken up, the fluid then proved effective. 

 Of course, I did not use it directly, but first filtered it five to six times 

 through triple filter paper, and then added the salts obtained by the 

 evaporation of sea-water so as to make it of the same concentration 

 as that of normal sea- water (about 4 per cent). When put into this 

 water, unfertilized eggs of both Sphaerechinus and Arbacia showed 

 signs of cleavage each, of course, only responding to the extract of 

 sperm of the same species. I may, however, remark at the same time 

 that not a very large number and by no means all of the eggs responded; 

 it was usually the case that the eggs of one individual would react to 

 one and the same sperm fluid, but not those of another. In the best 

 cases the segmentation proceeded regularly to the four-cell stage, but 

 afterward it became abnormal and the blastomeres which were very 

 dissimilar in size fell apart from one another, probably as the result of 

 the absence of the vitelline membrane. The velocity of development 

 was much slower in eggs so treated than in normally fertilized ones. 



I have no intention of criticizing Winkler's efforts; they 

 were certainly a step in the right direction. But it must be 

 pointed out that the experiments are not free from objection. 

 In the first place, his sea-water was made much more alkaline 

 than is normal, owing to the fact that by first evaporating it 

 he drove out the C0 2 and converted the bicarbonate into car- 

 bonate. He afterward restored the sea-water to its normal vol- 

 ume by adding distilled water. The increase of alkalinity thus 

 produced will alone lead to such results as Winkler describes, 

 as I showed years ago. 



For several years I have tried in vain to repeat Winkler's 

 experiments; results such as he described can be obtained 

 by using slightly hypertonic or hyperalkaline sea-water, or 

 the two together; but these results are also obtained when no 

 sperm is added to such sea-water. Before the appearance of 

 Winkler's paper I myself had examined the effect of various 

 enzymes upon the unfertilized sea-urchin egg, but with negative 

 results. At my suggestion, Professor W. J. Gies, of Columbia 

 University, undertook a series of experiments in which he 

 subjected the spermatozoa to every known method that leads 



