8 ON THE DIFFERENTIAL REACTION TO VITAL DYES EXHIBITED BY 



Protocol: Rat 333, injected intraperitoneally with fresh aqueous solutions of 0.5 per cent 

 trypan blue, February 21 to 24, inclusive, 2 c. c. each day. 



February 26: Animal stained a deep blue. Subcutaneous tissue deep blue. Under the low- 

 power an elegant distinction of cell types is evident. 



The oil shows this to be founded on the smaller, sparser, punctate, and linear nature of the 

 fibroblast deposits in contrast to the macrophages, which are here characterized by fairly uniform- 

 sized (somewhat larger than mast-cell granules) vacuoles which fill the protoplasm, though not so 

 densely crowded as is often the case, and by a considerable number of somewhat smaller, refractive, 

 fat droplets (fig. 2). Neutral red (1:1000) gives its specific reaction, and under the low-power 

 marks the macrophages out with great emphasis owing to the accentuation of color. Janus green 

 (1:10,000) did not stain the mitochondria of the macrophages; 1:5,000 (exposed 1 minute) finally 

 showed a specific reaction in a few macrophages. Short, green, curved threads were seen lying 

 between the vacuoles which were slightly pink from the janus. 



The fibroblasts contain fairly numerous minute dye deposits. Some are granular, but the 

 great majority are linear, or elongated triangles. Neutral red superimposes itself on these vital-dye 

 deposits and, in particular, colors electively the linear endings of vacuoles, or the free linear struc- 

 tures, in the protoplasm. Janus green (1 : 10,000), exposed 10 minutes, stains typical mitochondria, 

 but since the morphology, and especially the color, of the fainter, vital-dye deposits is so nearly 

 identical with these, the distinction is well-nigh impossible, and one's belief in the success of the 

 reaction is dependent upon the sudden and evident increase in the number of bodies having mito- 

 chondrial characteristics after the application of janus to the cell. 



By the discovery of dyes which in fibroblasts, at variance with the description 

 which we have just given, establish a typical vacuolar segregation-apparatus (figs. 

 44, 5) similar to that of macrophages, and, again, by the discovery of dyes and of 

 dosages which greatly exaggerate the filiform modification (figs. 7, 9, 11) of the 

 fibroblastic segregation-apparatus, we believe it has been possible to show points of 

 fundamental similarity in the reaction of these two cell types, as well as those 

 where characteristic differences are evident. Our line of argument will be facili- 

 tated by a preliminary analysis of the phenomena of "vital staining" as displayed 

 by the macrophage cells. 



It will be recalled that Ehrlich, to whom we are deeply indebted for the initia- 

 tion of the whole field of inquiry into the selective effects of dyestuffs, saw in vital 

 staining with the acid dyes an ocular demonstration of his chemoceptor theory in the 

 domain of the new science of chemical therapeutics. 1 It will suffice as an answer 

 to this conception for us to state that the chemoceptor theorj^ would appear to have 

 been adequately disproved by the above-mentioned experiments which one of us 

 has carried out over a considerable period of time in conjunction with W. Schule- 

 mann and in which several hundred compounds, whose approximate chemical 

 constitution was known, were tested for their biological effect. Some 260 of these 

 dyestuffs were known to us, as far as their chemical structure was concerned, with 

 all possible accuracy. Our studies showed the inadequacy of any such theory in 

 enabling us to predict either the spread of the dyes throughout the body (the macro- 

 scopic vital stain) or the exact type of reaction which would be produced with the 

 vitally stained cells. Indeed, our investigations indicated that various physical 

 characters of the dye solutions were the determining factors in their so-called 

 "staining" effects. 



1 Ehrlich, Paul, 1013. Address in Pathology on Cheniiotherapy, delivered before the Seventeenth International Congress 

 of Medicine, British Medical Journal, 1013, vol. 2, pp. 3D3-359: "I will mention here — in order only to indicate a few 

 examples — the intra-vitam staining of the nerve trunks by methylene blue, the staining of cell granules by neutral red, and 

 the distribution of isamine blue in the so-called pyroll blue cells so carefully and excellently investigated by Edwin Goldmann." 



