THE TWO GREAT GROUPS OF CONNECTIVE-TISSUE CELLS. 25 



20 days. One would predict that the trypan-blue deposits in the fibroblasts will eventually form 

 sturdy crystals betraying no sign of their origin from vacuoles. It is to be emphasized that the 

 belief that these have all arisen from vacuoles in which both dyes are present is due to the impossi- 

 bility of deciding whether a great many of the vacuoles have or have not a small amount of blue in 

 them, the morphology of the vacuolar system being the same whether blue is or is not present. 

 Janus green shows typical filiform mitochondria. 



The same animal was injected further with a mixture of equal parts of a 0.1 per cent 

 solution of T 148 and a 0.1 per cent solution of trypan blue, May 16, 18, 20, 22, 24, 26, 28, 30, to 

 June 1 to 17, inclusive, 1 c. c. each day; June 18, 0.6 c. c. ; June 19 to 29, inclusive, 1 c. c. each day. 



July 16: Films from left thigh examined. Animal is stained a light violet-color. Skin of left 

 thigh light blue. Under the low-power, minute deep blue, apparently crystalline deposits are seen 

 in some cells, persumably macrophages. 



The oil confirms this. Not much change in the cell picture recorded May 14 has resulted. The 

 macrophages still, as a rule, possess their trypan-blue deposits in the form of small, intravacuolar 

 crystals. Free needles of a larger size are present in many cells, especially along blood-vessels. 

 Neutral red shows that the small vacuoles are numerous in macrophages and the tendency to 

 neutral red concretions marked. 



Fibroblasts are packed with T 148 vacuoles and exhibit no departure from the condition on 

 May 14, except that it is usual to find one or two trypan-blue deposits of deep blue, either seed- 

 shaped or needle-like, and unrelated toT 148 deposits. These are apparently the "sturdy crystals" 

 looked for. It is to be emphasized that there is no noticeable increase in the number of T 148 

 bodies which have been chosen for trypan-blue deposits, i. e., half a dozen only house the blue dye, 

 but the blue is now in crystalline form within or along the edge of vacuoles and also apparently free. 



We have also carried out experiments quite the reverse of those just cited. We 

 have employed dyes which create a large vacuolar segregation-apparatus after the 

 preliminary production of small pure crystalline deposits with trypan-blue isomers. 

 While some few of the original crystalline deposits happen never, subsequently, 

 to be enveloped by the vacuoles created by the second dosage, most of them are 

 rapidly inclosed in this way and with continued further dosage with the second 

 dye are eventually dissolved in its vacuolar fluid. It could be expected, and, in- 

 deed, it has invariably resulted in all experiments of this type, that the main mass 

 of the large segregation-apparatus produced by the second dye contains no color 

 contamination with the dye first employed. A simple explanation for this is at 

 hand in the fact that most of these structures are de novo formations, originating 

 only after the inauguration of the second dye treatment. We have, in such ex- 

 periments, scanty "dye bodies " of pure color of the first dye, many of pure color of 

 the second vital dyestuff employed, and a considerable number of mixed "dye 

 bodies" mingled with them. (See figs. 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 

 61, 62, 63, 64.) 



Protocol: Rat 16, injected intraperitoneal^ with a 0.5 per cent solution of trypan blue, 

 January 30, February 2, 5, 8, 11, 14, 17, 20, 23, 26, March 1, 4, 10, 13, 16, 19, 22, 25, 

 and 28, 1 c. c. each day. 



March SO: Films from right thigh examined. Animal is stained a bright blue. Under the 

 low-power, deep-blue dye deposits are present in both types of cell. Some macrophages can be 

 distinguished by their larger dye deposits. Fibroblasts are seen to be heavily laden with deposits 

 to the full extent of their processes. 



Under the oil, the macrophages appear to have even less dye than the fibroblasts. Their dye 

 content varies from one or two blue vacuoles containing crystals to great numbers of such. Where 

 much blue dye is present deep blue crystals lie free in the cytoplasm between vacuoles (fig. 50). 

 Pale blue and colorless vacuoles are numerous in macrophages containing but little blue dye. 

 Neutral red gives a specific stain in the macrophages. Apparently pure red vacuoles are very 

 numerous in many of these cells, which show relatively little blue. The large vacuoles containing 

 blue crystals are stained by neutral red. Some blue crystals are present unaffected by the neutral 

 red and lie free in the cytoplasm. 



