28 BASIC METHODS FOR EXPERIMENTS ■ 



condition, for three chief reasons. First, they often crowd the 

 eggs in a small volume of sea-water. Eggs placed directly 

 from the ovaries in very little sea-water often fail to maturate; 

 this failure is an effect of CO2. Butyric acid and insemination 

 also inhibit maturation. On the other hand, maturating eggs 

 are highly susceptible to CO2, butyric acid, and elevation of 

 temperature because all of these agents initiate development. 

 Shaking matured eggs, as Mathews (1901) has shown, causes 

 them to develop. In this last case my own observations indicate 

 that CO2 here also plays a part. 



Secondly, workers, because they do not use sufficient care 

 in opening the animals and removing the gonads, too frequently 

 contaminate the eggs with perivisceral fluid or tissue extracts. 

 Asterias eggs are the most sensitive that I know. The worker 

 will obtain infinitely more constant results if he treats this egg 

 with respect. In addition, he would save time and would avoid 

 the needless destruction of animals. 



Thirdly, the practice of chopping up the ovaries for obtaining 

 eggs mitigates against securing a high per cent, of normal devel- 

 opment. By this method many young ovocytes are released 

 whose germinal vesicles are not stimulated to break-down when 

 the eggs are brought into sea-water. In this case, one may often 

 count more eggs with intact germinal vesicles than those whose 

 germinal vesicles are breaking down. 



The worker can prove to his own satisfaction that the 

 method for handling Asterias eggs which I have outlined above is 

 a good one. First, let him take shed eggs and inseminate them. 

 Next, inseminate eggs from the ovaries of the shedding female as 

 outlined above. Finally, let him now cut up the ovaries and 

 inseminate the eggs thus obtained. He will find that while the 

 eggs of the first and second lots are about the same, as they 

 reveal by their high per cent, and normality of development 

 through the bipinnaria stage, the eggs from the cut-up ovaries 

 are distinctly inferior in both respects. 



Asterias glacialis 



The method for Asterias forbesii and vulgaris, I have used 

 with success at Naples for A. glacialis. In January, 1929, I 

 obtained my first lot of shedding A. glacialis at Naples. 



