MITOCHONDRIA IN TISSUE CULTURES 377 



on the growth it was drawn off and the tissue again bathed in 

 a fresh drop of the warm solution free from dye. The mito- 

 chondria take up -the dye within a few minutes and remain 

 stained from thirty minutes to two hours. So far as we have 

 observed, the intensity with which the mitochondria stain does 

 not depend upon the strength of the solution. A very weak 

 solution (1-100,000) gives as intensely stained mitochondria as 

 does a strong solution (1-5000). A weak solution, such as 

 1-100,000 Janus green, stains only the mitochondria a blue- 

 green, while the cytoplasm, nucleus, and nucleolus remain clear. 

 A strong solution (1-5000), however, stains the cytoplasm a pale 

 green, the mitochondria a darker green, the nucleolus green, 

 and the nucleus a more or less violet-green. 



Nile blue B extra and hrilliant cresyl blue 2 b 



Aside from Janus green, no dye used in these observations 

 stained the mitochondria in the living cell. Both nile blue A 

 concentrated or B extra and brilliant cresyl blue 2 B, however, 

 did stain the mitochondria after the death of the cell, especially 

 after fixation either with neutralized formalin vapor or osmic 

 acid vapor. This is interesting in connection with the work 

 of Lorrain Smith ('08) on differential stains for fats. He states 

 as follows: 



It was observed that watery solutions of nile blue sulphate (A), a 

 colour stuff of the oxazine series, stains the fat globules contained in 

 tissue cells in various colours. In the majority of cases the fat globules 

 are stained a brilliant red; occasionally globules are present which take 

 a blue stain, and not infrequently the colour is due to a mixture of blue 



and red We may express the reaction in the following 



way: The fatty acid combines with the oxazine base to form a blue 

 soap, whereas both neutral fat and fatty acid merely dissolve the rela- 

 tively weak oxazone base (red). 



He remarks in relation to tissues fixed with formalin that 

 the globules stain readily either red or blue according to their 

 composition : 



When a globule contains a small amount of fatty acid and a large 

 amount of oxazone base is present in the solution of the dye, the globule 

 becomes predominately red, whereas if the stain is relatively weak in 

 oxazone the blue colour of the oxazine staining is more apparent. 



