480 
Notes, 
and female plants were kept in separate dishes, and were covered 
over so as to prevent drying up. This method gave far better results 
than those more usually advocated. On the appearance of the 
extruded sexual products, the female receptacles were placed in sea- 
water, and after the complete liberation of the oospheres, a few male 
branches with ripe antherozoids were first placed in a capsule of sea- 
water until it became turbid owing to their number. If on examination 
the antherozoids proved to be active, small quantities were added to 
the vessels containing the oospheres. The latter were then fixed at 
intervals of five minutes during the first hour, and then at intervals 
of fifteen minutes, up to six hours after the addition of the anther- 
ozoids. After that, samples were killed at longer intervals up to 
three days, and this was continued till we had material fixed at all 
stages for the first fortnight. At first we used sea-water in which to 
keep the embryos growing, but a proper solution of Tidman’s sea-salt 
was found to answer quite as well. 
For fixing, we tried the following reagents — chrome-alum, picric- 
alum, Mann’s picro-corrosive, corrosive sublimate, and acetic acid; 
these were all dissolved in sea-water ; absolute alcohol, Flemming s 
and Hermann’s solutions, and the vapour of osmic and formic acids. 
The Flemming’s (strong formula) and Hermann’s solutions were 
diluted with equal parts of sea- water. The first three fixatives were 
unsuccessful, acetic-corrosive yielded fair nuclear figures, but the 
material proved very brittle, and the spores were somewhat distorted. 
A portion of the cytoplasm was disorganised and the polar radiations 
were not preserved. Absolute alcohol fixed the oospheres and newly 
fertilised spores without distortion, but was useless for all other stages. 
Vapour fixing with osmic acid succeeded better than any of the 
preceding reagents, but was greatly inferior to either Hermann’s or 
Flemming’s solutions in preserving the protoplasmic structure in an 
unaltered state. 
After the material had been fixed it was dehydrated and passed in 
the usual way into paraffin, the temperature of which was not allowed 
to exceed 5o°C., and it was then cut with the microtome. The 
sections were stained with Fleidenhain’s iron-hsematoxylin, with Flem- 
ming’s triple stain, and a large number of other dyes. The results, 
which were compared carefully, led us to rely chiefly on the two 
staining processes mentioned, but at the same time we often obtained 
valuable preparations with other staining reagents as well. 
