10 E. V. COWDRY 



disc, each space on which had been estimated, with the com- 

 bination used (Zeiss apochromatic objective 1.5 mm. and com- 

 pensating ocular 4), to be equivalent to 1.5/x- Cells, the greatest 

 diameter of which measured 30^ or less, were classified as small, 

 the others being referred to as large. 



In the study of fresh tissues the greatest care was taken to 

 obtain isotonic media. Where possible the cells were observed 

 in their native tissue juices without the addition of any foreign 

 fluid. Occasionally, however, it was found necessary to employ 

 some sodium chloride. Ringer's or Locke's solution. I used janus 

 green (M. L. B.) and diethylsafranin, which I made myself 

 ('14 b) from diethylsafraninazodimethylanilin, extensively as 

 vital dyes for mitochondria; and nilblau B extra (B. A. S. F.), 

 methylene blue medicinale (M. L. B.) and new methylene blues 

 GB, N, NSS, NSSF, NX, R and RRR (Cassella), which Dr. 

 H. M. Evans kindly gave to me, for the lipoid globules and 

 Nissl substance. 



The standard mitochondrial methods of Bensley ('11, p. 309), 

 Altmann ('90, p. 27), Benda ('02, p. 752), Meves ('08, p. 832) 

 and Regaud ('11, p. 3) were employed in the study of fixed 

 tissues. The technique as originally outlined by these investi- 

 gators was adhered to with as little deviation as possible. The 

 temperature and time were not kept constant. Bergamot oil 

 was often substituted for xylol as a clearing agent, but only after 

 experimentation had shown that doing so did not alter the 

 specificity of the stain. The duration of impregnation in paraf- 

 fin at 60°C.=»= varied from one-half to two hours according to 

 the size of the piece of tissue. Solubilities were tested by fix- 

 ation in fluids containing different amounts of acetic acid, alco- 

 hol, corrosive sublimate and formalin, because it is well known 

 that these substances exercise a destructive influence on mito- 

 chondria. This was done by fixation in Zenker's fluid with and 

 without acetic acid, Carnoy's fluid, absolute alcohol and formalin. 



The observations were carefully controlled by comparing the 

 unstained cells with vitally stained ones and permanent prep- 

 arations. Where possible a number of animals of the same 

 species were studied in order to eliminate individual variations. 



