252 THE EMBRYOLOGY OF THE HONEY BEE 



which fixing fluid gives the truest pictures of these cells can how- 

 ever only be determined by careful comparison with living eggs. 



Picro- formal proved to be the best medium for fixing eggs in- 

 tended to be studied entire, and since the other media showed no 

 decided advantage in other respects, it was most frequently em- 

 ployed. Its reason for superiority appears to be due to the 

 circumstance that it does not coagulate the yolk spheres, conse- 

 quently the yolk remains clear and transparent. Embryos fixed 

 in this fluid and mounted entire have a brilliancy and transparency 

 not seen in specimens fixed in other fluids. 



The eggs were in all cases preserved in eighty per cent alcohol. 



Of the five fixing fluids used, Gilson's and Petrunkewitsch's 

 fluids harden most, eggs fixed in these media becoming rather 

 brittle, while acetic alcohol hardens the least. Eggs treated with 

 this mixture are quite elastic and may be manipulated without 

 much danger of breaking, although they are rather easily distort- 

 ed. This is true also of eggs treated with picro-formol, although 

 not to the same extent. 



The chorion of the egg of the bee, although excessively thin, is 

 nevertheless relatively impermeable, especially to aqueous fluids. 

 After fixing and hardening, and before staining or clearing is 

 attempted it is therefore necessary to puncture the chorion, which 

 lies close to the egg except at its two poles. The chorion may be 

 punctured or ruptured here, but the most satisfactory solution of 

 the difficulty is to puncture the egg in one or two different places 

 with a fine cambric needle, rubbed on an oil stone to an extremely 

 fine point. This is best done by watching the progress of the 

 sharpening under a binocular microscope. The punctures were 

 usually made along the dorsal mid-line of the egg since this region 

 is not occupied by the embryo until near the close of development. 



Mayer's carmalum was used for staining eggs intended to be 

 mounted entire. The eggs were stained from two to forty hours, 

 and thoroughly destained in acid alcohol. This method yielded 

 uniformly good results. Iron haematoxylin was found most sat- 

 isfactory for sections, used with or without a counter stain. 

 Mayer's paracarmine (alcoholic) was used for staining eggs pre- 

 liminary to embedding. 



It is absolutely necessary that clearing be accomplished grad- 



