ON EGGS OF MARINE ANIMALS 49 



Acting on echinid eggs, weak hypertonic sea-water made up 

 by adding 8 cc. of 2.5M NaCl to 50 cc. of sea-water, as originally 

 used by Loeb, calls forth development without well separated 

 membranes, with poor cleavage and with swimming forms that 

 do not rise to the surface of the water. Again, unless the 

 worker intends frankly to study this phenomenon, he would 

 do well not to use this method. This weak hypertonic sea-water 

 is used best in combination with butyric acid, as given beyond. 



Hypertonic sea-water calls forth development closely simu- 

 lating the normal in several species of eggs. Among these are 

 eggs of molluscs, worms, the starfish and echinids. Since the 

 echinid egg is the one most extensively employed in the study of 

 experimental parthenogenesis, I give the methods which I have 

 found to be the best for inducing full development in it 

 (1922). 



From a single female take a thick drop of eggs known from 

 previous trial inseminations to be in optimum physiological con- 

 dition. Flood with 10 cc. of 2i^M NaCl or KCl in a watch- 

 glass mounted under low power of the microscope. Note 

 exactly with the stop watch the time to complete membrane- 

 separation. In the majority of cases one will find that 40 to 

 60 seconds exposure is sufhcient. Remove eggs to 250 cc. of 

 normal sea-water. With this method, the eggs should cleave 

 and 90 per cent, should develop into top-swimming plutei. 



Hypertonic sea-water may also be used by making up sea- 

 water plus 2i^M NaCl or KCl in the following proportions: 

 the salt solution in a series beginning with 95 parts and decreas- 

 ing by grades of 5 parts to 25 parts plus the sea-water beginning 

 with 5 parts and increasing in the same way to 75 parts respec- 

 tively. In these solutions, the duration of the effective exposure 

 increases with the amount of sea-water employed. All of these 

 concentrations except the last named require quick manipulation 

 on the part of the observer. Hence, he may prefer to use the 

 following: 



To a drop of eggs in optimum physiological condition is 

 added hypertonic sea-water made up of 80, 78 or 76 parts of 

 sea-water plus 20, 22 or 24 parts respectively of 2'^^M NaCl or 

 KCl. Under the low power of the microscope the time to 

 membrane-separation is noted and the eggs are removed there- 



