64 
R. KOCH. 
in Canada balsam. For the examination of the sputum with reference to tuber¬ 
culous bacilli, the second coloring can as a rule be omitted, so that the microsco¬ 
pic examination of such sputum preparation follows immediately upon the 
treatment of the same with nitric acid and alcohol. 
For the sake of a general view, I will recapitulate briefly the whole coloring 
process : covering glass preparations dried in the thinnest possible section, after 
the drying, three times heated in the flame; section preparations of objects, 
which are well hardened in alcohol; coloring of a solution consisting of 100 ccm. 
of aniline water, 11 ccm. of alcoholic methyl violet solution or fuchsine, 10 
ccm of absolute alcohol ; the preparations remain in the color solution at least 
twelve hours (the coloring of the covering glasses can be shortened by warm¬ 
ing of the solution) ; treatment of the preparations with diluted (1:3) nitric acid 
for some seconds ; washing in 60 per cent, alcohol for several minutes; (for cov¬ 
ering glasses, moving back and forth in alcohol several times is sufficient); second 
coloring in diluted vesuvian solution or metliyline blue for several minutes ; wash¬ 
ing again in 60 per cent, alcohol, getting rid of water by means of absolute 
alcohol, brightening in cedar oil; microscopic examination of the preparation ; 
enclosing of the preparation in Canada balsam if the same is to be preserved. 
As to the microscopic examination of the objects prepared in this manner, 
all that I have said in other works* about objects colored for the microscope, holds 
true also for these. In this case also structural relations, which make themselves 
manifest by the varying refractive power of the single parts of the tissue, are not 
to be settled ; it concerns us only to see the various color relations of the micro¬ 
scopic objects, that is to say, representations of absorption, as clearly and sharply 
as possible. The structural image whose effect is only disturbing must therefore 
be gotten rid of, which, as I have shown, can be done most completely with the 
help of the well-known illuminating apparatus of Abbe. The peculiar illumina¬ 
tion which this apparatus affords when it is used without “ abblendung,” cannot be 
borne by all systems of lenses. The last must be constructed with special refer¬ 
ence to this method of illumination. The greater an opening angle a system 
possesses, the better it is adapted for the observation of the images of absorption, 
with help of Abbe’s illuminating apparatus. For this reason oil-immersion sys¬ 
tems can accomplish the most in the investigation of colored objects. 
The covering-glass preparations, if rightly prepared, must possess so little 
thickness that the structure is formed of a single layer of objects, and in and of 
itself is little to be considered. These preparations can on this account be exam¬ 
ined simply in water and in case of need, a system of water-immersion is sufficient 
for them, if the field of vision be sufficiently brightened by a condenser. In the 
case of section-preparations, it is, on the contrary, impossible to set aside the 
structure formed by so many layers of tissue one above the other, unless the prep¬ 
aration is laid in a liquid which has high power of refraction. It is necessary 
to do away with the differences in refraction of the tissue, and to use the full 
illuminating power of Abbe’s apparatus, and must use its full power to the best 
advantage through the large opening angle of an oil-system. One may easily con- 
*Untersucliungen ueber dis Aetiologie der Wundinfectiouskrankheiten. Leipzig 1878, 
p. 31, etc. 
Mittbeilunengen aus dem Kaiserlicben Gesundbeitsamte. 1881, Vol. 1, p. 9. 
