vii ANNELIDA 131 



will immediately be shed, and in this way a natural fertilization of 

 the eggs is accomplished. 



The Trochophore issues on the second day and rises to the top of the 

 water. It can be reared through its entire development by supplying 

 it with a pure culture of the diatom Nitschia. Such pure cultures 

 can be obtained from Dr. Allen, Director of the Marine Biological 

 Station at Plymouth, and they serve as pabulum for many different 

 kinds of larvae. 



Pure diatom cultures were obtained originally by isolating under 

 the microscope a single individual of the species of diatom desired, 

 and then transferring it to a flask of sterilized and filtered sea- water. 

 The sea-water is first shaken up with animal charcoal and decanted 

 in order to remove all soluble toxins, and then passed through a 

 Berkfeldt stone filter, which removes all organisms, even bacteria. 

 To the sea-water is now added a certain amount of Miguel's solution, 

 about 2 drops per 100 c.c. of water, and the flask is stopped by a plug 

 of sterilized cotton wool. In a month's time a copious growth of the 

 desired diatom is obtained. 



If a pipette-full of such a culture be added to an evaporating dish 

 containing the larvae of Pomatoceros, these will develop normally 

 and eventually metamorphose into the adult worms, which form 

 tubes and attach themselves to the sides of the glass. In this way 

 the whole life-cycle can be controlled, and such larvae can be 

 examined living, or mounted whole, or examined by sections. 



The fixative found best is Eisig's mixture (see ante). The methods 

 of orientating, embedding, and cutting have been 1 fully described 

 in Chapter II. 



POLYGOKDIUS. CELL-LINEAGE 



Returning now to Polygordius we should remind the student that 

 this is a minute worm which burrows in mud and sand. The eggs are 

 excessively minute and very transparent, and the segmentation is 

 remarkable for its extreme regularity. The eggs are dehisced into the 

 sea by the breaking up of the parent's body and are fertilized there. 



Up to the 64-cell stage all the cells divide at the same time, so 

 that we have successive " cleavages " which successively divide the egg 

 into 2, 4, 8, 16, 32, and 64 cells, that, is six cleavages in all. More- 

 over a 128-cell stage is very nearly realized, for all the cells of the 

 64-cell stage divide nearly synchronously, except those forming 

 the prototroch and a few others of the upper hemisphere, which, 

 having reached the summit of their development, divide no more. 

 The macromeres are all precisely equal in size ; it is therefore at first 

 impossible to discriminate an A from a B, a C, or a D (see Chapter V.), 

 but in the later cleavage stages this can be done, owing to the different 

 way in which members of the second and third quartettes of 

 micromeres, given off from the different macromeres, behave. 



Shortly after the 64-cell stage has been reached cilia appear 

 on the cells destined to form the prototroch (Fig. 101, B). The 



