ROTATORIA OF THE UNITED STATES. 
277 
METHODS. 
The use of preserved mounted specimens has been the basis for the present work. 
It is only through the methods devised within the last decade by Mr. Charles F. 
Rousselet, of London, England, that the use of such preserved material has become 
possible in the study of the Rotatoria. Hence the complete lack hitherto of type or 
reference specimens among these animals. This has been one of the prime causes 
of the great confusion in the classification of the Rotatoria. A few of the genera 
have been worked over in the last few years with the use of preserved specimens by 
Mr. Rousselet and his collaborators in England. It is not too much to say that it 
will be necessary to *go over the entire group of Rotatoria in the same manner before 
order can be brought out of the present confusion. 
Killing and preservation . — The collections of preserved material on which the 
following paper is based were made as follows: The Rotifera were taken in various 
ways— by towing with the tow net in water free from vegetation, by washing aquatic 
plants in jars of water, by bringing into the laboratory quantities of aquatic plants 
together with some of the water about them, etc. Most of the Rotatoria come after 
a time to the lighted side of the vessel in which the material collected is placed. 
These are transferred in large numbers to a watch glass and placed beneath a 
simple microscope or low power of the compound microscope, where the movements 
of the organisms can be observed. 
Now a considerable quantity of Rousselet’s narcotizing fluid is mixed with the 
water in the watch glass. One-fourth as much narcotizing fluid as there is w'ater, 
or a larger or smaller proportion, may be used, as seems desirable from observation 
of the movements of the animals. Rousselet’s narcotizing fluid consists of 2 per cent 
solution of hydrochlorate of cocaine, 3 parts; methyl alcohol, 1 part; water, 6 parts. 
This causes the animals to swim slowly and gradually to settle to the bottom; they 
will soon die, and if allowed to die unfixed will be quite worthless for study, destruc- 
tive changes taking place in the tissues at the moment of death or perhaps even before. 
As soon, therefore, as most of the rotifers have sunk to the bottom, as much of the 
water as possible is drawn off from above them with a pipette. Then a small amount 
of 0.25 per cent osmic acid is introduced, which kills and fixes the rotifers at once. 
Now remove the osmic acid as quickly as this can be done without taking up too 
many of the rotifers (within a minute or two if possible), and wash several times 
in distilled water. In thus fixing the rotifers in large numbers at once, it is usually 
impossible to draw off the osrhic acid as soon as would be best, so that the animals 
become much blackened. But the blackening may be removed later with hydrogen 
peroxide. If the osmic acid has been used at the right time usually a majority, or at 
least many, of the rotifers will lie found to be fixed well extended. But as the time 
required for narcotization varies with different species as well as with different 
individuals of the same species, many of the animals will be found contracted or 
with the structure partly obscured by degenerative changes. With practice, how- 
ever, it will become possible to secure a sufficiently large percentage killed in good 
condition to make the collection very valuable. 
For study of the loricate Rotifera it is advisable to kill some part of every col- 
lection directly by means of osmic acid, without previous narcotization, for in the 
loricate rotifers some of the most important distinctive characters can best be seen 
in contracted specimens. 
