^ETIOLOGY OF TUBERCULOSIS. 
63 
ing unaffected by nitric acid, is soluble in alcohol from sixty to seventy per cent, 
if the preparations are put immediately from the acid into the alcohol. A longer 
remaining of the preparations in alcohol appears to make the coloring matter 
finally insoluble also for alcohol, and it is therefore to the purpose not to wash 
the preparations in water after their treatment with nitric acid, but to place them 
directly into the alcohol. 
The method of uncoloring followed by me is as follows : By the help of a 
platinum wire, which is melted into a little glass staff, the preparations are lifted 
out of the color-solution and laid into nitric acid diluted with three to four parts 
of water. In this they are moved back and forth for some seconds, until they 
have taken a greenish blue color, and are then put directly into a vessel with 60 
per cent, alcohol. In the alcohol they remain only about ten to fifteen minutes, 
after which they receive the after-coloring now to be described. 
In preparations treated with nitric acid and alcohol the component parts of 
the tissue are wholly colorless, or possess only a slight bluish tone of color, while 
the tuberculous bacilli have retained an intense blue color. Relative posi¬ 
tions of the bacilli to their surroundings, are, owing to the nature of the prepar¬ 
ations, difficult of proof. It is also very difficult to find single bacilli in the 
tissue, whose structure is made as good as completely invisible by the peculiar 
method of illumination, which will be described later, and for this cause it is 
necessary to give to the tissues a coloring of the nucleus. In order to obtain as 
striking a contrast as possible, between the coloring of the bacilli and the cell- 
grains, a yellow or light brown is chosen for the supplementary coloring material, 
when the bacilli are blue; a green or blue is chosen when they are red. For the 
first case vesuvian is best adapted, for the second methyline blue. Both coloring 
materials must nevertheless be used only in weak solutions, and not for too long 
a time, in order that just sufficient coloring of the grains may be obtained, lest 
single bacilli be hidden by too darkly dyed masses of grains. I use for the sec¬ 
ond coloring a watery, freshly filtered vesuvian-solution, which to a depth of 2 
ccm. is just barely transparent. On this the uncolored covering-glass preparations 
are so laid that they float with the prepared section downwards. Section prepar¬ 
ations remain in it some minutes. It is not necessary that the section prepara¬ 
tions, when they are brought from the alcohol into the vesuvian solution, should 
be completely colorless, because they must later be again treated with alcohol, in 
order to get rid of the water in them, and will then lose whatever blue coloring 
matter will have remained. 
One takes the preparations out of the vesuvian solution aud puts them again 
into ten per cent alcohol and out of this into absolute alcohol. The further treat¬ 
ment is the familiar one, only it is to be recommended that for brightening the 
preparation, instead of oil of cloves, oil of turpentine, or still better, cedar-oil be 
used, as these do not draw the aniline out of the preparations. With reference 
to enclosing them with Canada balsam, I would say that a balsam diluted with 
oil of turpentine appears to be the best adapted. Very thick balsam, which must 
be warmed in order to enclose the preparation, must not be used, because in 
warming, the tuberculous bacilli usually lose their color quickly. 
Covering glass preparations can be examined immediately after the washing 
off of the vesuvian solution with water, or they can be dried again and enclosed 
