882 E. J. ALLEN AMj E. W. NELSON. 



Methods. — In carrying out the experiments to be described 

 in this section the procedure has been as follows: All media 

 have been prepared from sterile sea-water, and sterile vessels 

 and instruments have always been used. The cultures have 

 usually been made in 60 c.c. of liquid, in short-necked, wide- 

 mouthed flasks of 125 c.c. capacity. When a number of 

 cultures were to be compared, the flasks were kept standiijg 

 in a row together in such a way as to keep the physical 

 conditions as similar as possible. Control cultures in standard 

 media were included in each series, so that results from 

 different series could be compared by reference to the 

 controls. The various media were inoculated from a persistent 

 culture of a species of plankton diatom, which in the great 

 majority of cases was Thalassiosira decipiens (p. 412). 

 When preparing the different media the methods used were, 

 as far as possible, identical, and although only about 60 c.c. 

 was needed for a culture, a litre was made up, so that errors 

 due to measuring very minute quantities might be avoided. 

 The media were all freshly prepared for each compaiative series 

 of cultures, the same sample of sea-water being used, when 

 the basis of any two or more was the same. Comparative 

 estimates of the amount of growth in the different cultures 

 were made by eye alone. Any difference between amounts of 

 growth that has been described here as appreciable has always 

 been accompanied by a marked difference in appearance to 

 the eye on holding the cultui-es up to the light. A few drops 

 from each culture were also, from time to time, examined micro- 

 scopically, as a test of the quality and purity of the growth. 



The Sea-water Employed. — The sea-water employed as 

 a basis for the culture media has been either (1) "outside 

 water " or (2) " tank-water." A general description of these 

 will be found on pp. 372-374. An accurate chemical analysis of 

 both ty{)es of water would probably make clear many difficult 

 points, but, as already pointed out, no chemical methods of 

 sufficient delicacy have yet been devised. 



We have seen that if we compare '^tank-water," i.e. water 

 from the closed circulation of the Plymouth Aquarium, with 



