THE TWO GREAT GROUPS OF CONNECTIVE-TISSUE CELLS. 21 



addition, at various points, either within the vacuoles or between them, crystalline 

 condensation of the vital dye at length occurs. The "saturation crystals" may 

 be produced typically in the case of vital dyes with which it is impossible to pro- 

 duce crystals with low dosages of the dye. In the case of the macrophages of 

 animals treated with dye 245, the "saturation crystals" appear to "drain" the 

 vacuoles of the entire segregation-apparatus of these overloaded cells, which are 

 now colorless, and between which the characteristic long, deep-saffron needles of 

 the vital dye appear. (See figs. 42, 43.) 



Protocol: Rat 2286, injected intraperitoneally with 1 per cent solution of dye 245 (an 

 isomer of vital new red) with the following formula : 



= N-{ Vnh — c— HN-( Yn = 

 /H > ' « N { H\ 



May 16, 17, 19, 22, 25, 28, 31, June 3, 6, and 9, 1 c. c. each day; June 12, 0.8 c c. 



June 13: The animal is stained a brilliant red with some orange hue. Subcutaneous tissue of 

 thigh is a deep fiery red, and even thin films are bright pink. Under the low-power one detects 

 immediately macrophages as faint-pink masses containing deep-red linear deposits which, even at 

 this power, are appreciated as crystals. One can not with certainty make out the fibroblasts. 



The oil shows that the macrophages are entirely full of small vacuoles in which the color is 

 extremely pale. Among these are straight, sharp, needle-like, deep eosin-colored crystals. These 

 form complicated "brush-piles" in the case of macrophages along blood-vessels and are present to 

 some extent in practically all of the macrophages (fig. 42). 



The fibroblasts and their processes are full of fairly large, irregular-shaped, faint orange or 

 yellow colored vacuoles. A small portion of these vacuoles have red crystals and needles related to 

 them (fig. 43). 



Still another set of circumstances may give a deposition of vital-dye crystals 

 within these cells. We refer to the appearance of crystals in the case of the em- 

 ployment of a dye and a dosage which produces a vacuolar segregation-apparatus 

 devoid of these structures, but which in decolorizing tends to concentrate its dye- 

 content between the vacuoles in crystalline form. (Note the appearance of decol- 

 orization crystals in the case of dye T 148, rat 18-2, page 13 and rat 19-2, page 22, 

 which appeared one month after dosage was stopped.) 



4. Although Kiyono (1914) and Pappenheim and Nakano (1913) have reported 

 "double-staining" experiments with acid and basic dyes, or with two acid dyes, 

 it would appear that no one has made cytological studies of great accuracy in this 

 field. In particular, the necessary accurate records on the actual effect of the first 

 dye have not usually been secured from a study of skin samples taken under an- 

 esthesia before dosage with the second color is begun. The two vital dyes employed 

 for such experiments must differ as widely as possible in their color, a crimson-red 

 and a deep-blue dye giving admirable distinction. In two dyes which, when ad- 

 ministered alone, produce approximately the same effect, i. e., enter the cell at the 

 same rate and are segregated in the same way, pronounced color mixture in the 

 segregation-apparatus always occurs after dosage with the second compound. It 

 is true that some vacuoles appear to carry only the first color and others clearly 

 only the second, but deposits of uncontaminated color are rare. Such experiments 



