and Laboratory Methods. lOlo 



NOTES ON TECHNIQUE, ML- 



L The Preparation of Sections of Hydra for Class Work. 



The preparation of satisfactory sections of Hydra for microscopical study of 

 the tissues has always been attended with more or less difficulty. Usually the 

 cell boundaries are poorly defined, sub-epithelial cells are not clearly marked ofif, 

 and on account of the presence of large quantities of metabolic products — fat 

 globules, etc. — in the cells, the sections as a whole have an indefinite, crowded 

 appearance. After some experimentation the following method of preparing and 

 staining sections of Hydra was tried with excellent success. Its results are very 

 satisfactory ; cell boundaries are clearly marked, so that sub-epithelial cells may 

 readily be seen by the student, the muscle processes appear with entire distinct- 

 ness, and finally the cytological relations of the cells are well defined. 



The method used is in detail as follows : Large specimens of the common 

 brown Hydra (H. fusca) are placed in Stender dishes filled with filtered tap water 

 and allowed to "starve." The purpose of this " starving" process is to rid the 

 cells of stored-up fat and other metabolic products. Since the animals are placed 

 in dishes containing only clear water they are unable to feed and so use up grad- 

 ually these reserve and intermediate products of assimilation. This " starving " 

 is perhaps the most important step in the process, as on its thoroughness the 

 character of the sections depends. The length of time which must be allowed 

 for the animals to become entirely free of fat globules will depend on their con- 

 dition at the beginning. We have found in practice that from one to two weeks 

 is usually a sufiicient length of time. When the process is completed the ani- 

 mals are very light, in fact almost white, in color and are more than ordinarily 

 transparent when fully extended. Some of the specimens may die in the course 

 of the " starving" process, but not any considerable number are lost in this way. 



When thoroughly •' starved " the animals are fixed in vom Rath's picro-osmo- 

 aceto platinic chloride mixture^ for about an hour, then washed in methyl alco- 

 hol, blackened for several hours in pyroligneous acid, again washed in methyl 

 alcohol, removed to 70 per cent, alcohol and carried up through 90 per cent, 

 and absolute alcohol, cleared in cedar oil, and embedded in parafiin in the ordi- 

 nary way. 



The animals are killed almost completely extended in the following way. A 

 Hydra isolated in a pipette with a small amount of water is placed on a slide or 



* These " Notes " do not make any claim for originality, but are merely detailed accounts of 

 some applications of ordinary methods which have been found in practice to accompHsh i.i a 

 satisfactory way the desired ends. They are offered in the hope that they may prove useful to 

 teachers. R. P- 



1 vom Rath, O. Zur Conservirungstechnik. Anat. Anz. II : 280-288, .1895. 

 Formula (p. 283) : 



200 c. cm. saturated aqueous solution of picric acid. 

 Added in ( 25 c. cm. 2 per cent, aqueous solution of osmic acid, 

 the order ] i g. platinic chloride, dissolved in 10 c. cm. of water, 

 given. r 2 c. cm. glacial acetic acid. 



Fixation in Hermann's fluid, with subsequent treatment with pyroligneous acid in the same 

 way, has been found to give equally good results. 



