TECHNIQUE. 



transferred to weak glycerine, where dissections were made with No. 12 Sharp's 

 cambric needles under a high power binocular microscope. When mounted in 

 weak glycerine the embryos could be rolled in various directions without injury. 

 For permanent preparations the specimens were dehydrated through the alcohols, 

 cleared in equal parts of absolute alcohol and cedar oil for 12 hours, cedar oil 

 for Ci hours, and xylol for 1 hour. Then they were mounted in balsam. 



Musculature. For a study of the nymphal musculature, dissections of 

 newly molted specimens were made in glycerine. In toto preparations were 

 made by killing the specimens in formalin, dehydrating through the alcohols, 

 clearing in xylol, and mounting in balsam. Serial sections were cut 10 microns 

 in thickness, and stained with Delafield's hematoxylin and picro-fuchsin. 



External Morphology. Specimens used for study of the external mor- 

 phology were boiled in 5% KOH until only the chitinized parts were left. Then 

 they were stained with picro-carmine. 



Internal Metamorphosis. The material used for the internal meta- 

 morphosis of the mouth-parts was Hexagenia recurvata. 



For histological examination, specimens were treated as follows: 



Hot water 1 minute 



! felly's fluid 12 hours 



Running water 12 hours 



67% alcohol 24 hours 



82' I + tincture of iodin 72 hours 



90' - alcohol 12 hours 



95% alcohol 12 hours 



Abs. alcohol 12 hours 



Xylol 6 hours 



Xylol-paraftin 6 hours 



Pure paraffin 3 hour^ 



Sections were cut 9 microns thick, and stained from water with Giemsa's blood 

 stain (1 drop: 1 ce. H,0) for 45 minutes. They were washed in distilled water 

 until a faint pinkish tinge appeared ; then dehydrated through the alcohols, cleared 

 in cedar oil, passed through xylol, and mounted in neutral balsam. Material 

 fixed in Zenker's fluid and stained with Delafield's hematoxylin and eosin gave 

 good results for muscle degeneration, but did not differentiate the plasma cells. 



Plasma Smears. For plasma smears from nymph, subimago and imago, 

 a thoracic leg was dipped in 95% alcohol, allowed to dry, and then severed. The 



*Hel.ly's Fluid: Potassium dichromate, 2.5 gms. ; sodium sulphate, 1 gm. ; mercuric- 

 chloride, 3 gms.; .6% sodium chloride solution, 100 cc. ; formalin, 5 cc. added just before 

 using. This fluid preserves the granules in the plasma cells and permits their identification 

 in sections. 



