The Idcntijicatioti of Intracellular Structures. 103 



may go quite black in osmium tetroxide solution. This has been 

 interpreted to mean that apart from any fat content of the lecithin 

 molecule itself, there is a subsequent development of other free 

 fats, among which is constantly found olein. Formalin used as a 

 fixative is found to preserve yolk-discs, but in the case of Apanteles, 

 a Bracon fly, I find that such fixation does not prevent the discs 

 from shrinking during passage through up-graded alcohols, while 

 fixation in chrome-osmium mixture (e.g. Flemming's modified 

 formula) prevents shrinkage. This I consider shows that some 

 part of the insect yolk is not fixed by formalin, and is left soluble 

 in alcohol and subsequently washed out. Whether proteid, fat, or 

 cholesterin, this soluble matter is undoubtedly present ; I consider 

 that it is a fat. In Amphibia, and possibly in most other animals, 

 the older yolk-discs stain most intensely in iron-h?ematoxylin, 

 and in some forms Benda's stain (crystal-violet following alizarin) 

 strongly colours yolk-discs of all sizes a deep violet ; these results 

 are possibly traceable to a staining of some proteid (albuminous 

 matter) contained in the disc in addition to the lecithin, for I find 

 in the ovotestis of the snail that the young yolk-discs which 

 become yellowish in osmic acid refuse to stain either in iron- 

 hffimatoxylin, Altmann or Benda. Under this head the conclu- 

 sions I have come to by means of microchemical tests are as 

 follows : — 



1. The yolk-discs when newly forming do not become black 

 with OSO4 solution, but stain a yellowish colour somewhat deeper 

 than the general yellow colour of the ground cytoplasm ; as they 

 grow larger they become brown with osmic acid, and later may 

 become quite black. 



2. The so-called yolk- discs of some animals are, when full- 

 sized, formed of several categories of substances, of which lecithin, 

 proteid and fat are the most important components. 



3. The compound structure of the fully formed yolk-discs leads 

 to a variety of results under different techniques whose special 

 properties may be that of preserving either fat, proteid or lecithin 

 separately, or of preserving two or more together, and this result 

 is further confused by the fact that in yolk-discs of different 

 animals the quantities of the three above-mentioned substances 

 may vary. 



4. Unless care is taken, the fairly general association of lecithin 

 and free unsaturated fat leads to a confusion of fat with the yolk- 

 discs, for both may go black under treatment with osmic acid. 

 This difficulty is got over by dehydrating the material in acetone 

 instead of alcohols ; acetone dissolves away the fat and leaves the 

 lecithin ; or it may also be avoided by fixing in formalin and 

 dehydrating and clearing in alcohols and xylol, which dissolve fat 

 and leave yolk (lecithin). 



