106 DISTRIBUTION OF MITOCHONDRIA IN THE PLACENTA. 



method of Bensley, the copper-chrome-hematoxylin method of Bensley, the 

 fuchsin-picric acid method of Altmann, the alizarin-anilin-gentian violet method of 

 Benda, and the iron-hematoxylin method of Regaud. The best results were obtained 

 by fixing according to method IV B of Regaud and staining by the fuchsin-methyl 

 green method of Bensley. The great variation in the number and size of mito- 

 chondria renders differentiation by the iron hematoxylin method very difficult, as 

 the minute mitochondria in some cells are completely decolorized while adjacent 

 cells containing an abundance of large mitochondria remain deeply stained. 



In Regaud's method IV B the fixative is made up of neutral formalin 1 part, 3 

 per cent aqueous solution of potassium bichromate 4 parts. The formalin is neutral- 

 ized with magnesium carbonate. The tissues are fixed 4 days, the solution being 

 changed daily; they are then mordanted for 8 days in a 3 per cent aqueous solution 

 of potassium bichromate, washed 24 hours in running water, dehydrated in 50 

 per cent, 70 per cent, 95 per cent, and absolute alcohol, cleared in xylol and 

 imbedded in paraffine. 



Sections were cut 2 to 4 n in thickness and fixed to the slide by the water- 

 albumin method. The sections were freed from paraffine by xylol and passed 

 through graded alcohols to water, placed for half a minute in a 1 per cent aqueous 

 solution of potassium permanganate, then for half a minute in a 5 per cent aqueous 

 solution of oxalic acid. They were then washed thoroughly in water and stained by 

 Bensley 's acid fuchsin-methyl green method, which consists of the use of two stain- 

 ing solutions: i. e., (1) acid fuchsin, 20 gms.; anilin water, 100 c. c.; (2) 1 per cent 

 aqueous solution of methyl green. In the use of this method the following procedure 

 is to be carried out : Pour the acid fuchsin solution (a comparatively fresh solution 

 must be used) on the slide and heat the slide over a flame until the solution begins to 

 steam. Allow the stain to act about 6 to 8 minutes, heating 3 or 4 times to the 

 steaming-point. Pour off the stain, blot, and then wash the slide thoroughly in dis- 

 tilled water. Blot and pour 1 per cent methyl green on the slide. This must be 

 given time to stain the nuclei and protoplasm of the cells and is usually allowed to 

 act about half a minute. Then blot and dehydrate in absolute alcohol, clear in 

 xylol, and mount in balsam. If the sections are stained too deeply with methyl 

 green the minute mitochondria are obscured. If such sections are dipped for an 

 instant in 95 per cent alcohol part of the green is taken out and good differentiation 

 can be obtained. 



With this method the mitochondria are stained a bright red by the fuchsin and 

 the protoplasm is light green or unstained. The chromatin of the nucleus stains 

 green and the nucleoli either red or green. Unless otherwise stated, the following 

 descriptions are based upon sections prepared by the above technique. 



PLACENTA OF THE PIG. 



In the pig the chorion is covered by a single layer of cuboidal or columnar cells 

 which is applied directly to the uterine mucosa. A thin layer of structureless 

 substance, the so-called embryotrophe, which stains light green, lies between the two 

 tissues. The chorionic epithelium and uterine mucosa are thrown into inter- 



