74 \\VMAX REED GREEN. 



luced in the latter part of December, 1913. in laboratory 

 cultures, were subjected to the following conditions: One lot of 

 ,- \\as subjected to a freezing mixture for eight hours, one lot 

 of 6 to a freezing mixture for eight hours on two successive days, 

 and another lot of 6 were given the same treatment on three 

 successive days. In each case the eggs were transferred from 

 the freezing mixture to water at room temperature. The same 

 kind of treatment was applied to large numbers of eggs which 

 had been collected by scraping the surface of the ice on fresh 

 water ponds near Gary, Ind.. permitting the eggs to dry between 

 the freezings. Others were given the same treatment and then 

 dried for three months before being transferred to water at 14 C., 

 but all to no avail. 



Experiment 5. During the course of my work I have made 

 numerous attempts to induce development by clipping off the 

 small ends of the ephippia with scissors or scalpel, by pricking 

 the shell with a needle, and by dissecting it completely off. It 

 is hardly to be supposed that such treatment would have any 

 effect upon eggs that had never been dried, since they often have 

 the ventral edges of the ephippia only lightly apposed, it being 

 possible in some instances to look between them upon the egg- 

 inside, but conceivably desiccated eggs might be thus influenced. 

 It is quite a simple matter to spread the valves and set the egg 

 free. I have succeeded in three instances in placing ephippial 

 eggs with the shells removed in the brood pouch of asexual fe- 

 males. This did not incite development. 



On August 14, 1916, I .dissected the shells from 18 ephippia] 

 eggs and pricked them with an extremely fine glass needle. In 

 some cases I barely pierced the vitelline membrane, and in others 

 pushed the needle about one third tin way through the egg. 

 The membrane is quite tough and in most cases is under con- 

 siderable tension due to osmotic pressure. There is a quantity 

 of liquid underlying the membrane and minute quantities oi 

 this can be seen to exude I mm the wound in all cases, and it 

 the wound is made with too large a needle or too deeply the 

 :iular cytoplasm is extruded. Presumably such eggs are 

 imyed. Fourteen of these eggs seemed i< have been suc- 



v-fully treated and were placed in a slender dish with enough 



