Section Cutting and Staining: A practical introduction to histological methods for students and practitionersColman, Walter S.
Science
Section Cutting and Staining: A practical introduction to histological methods for students and practitioners
Colman, Walter S.
Microscopy
Much practice is required in using either of the methods before one can
judge accurately how long to leave sections in the staining reagents or
decolourising agents, and the beginner must not be discouraged if at
first he is unable to obtain good results although he follows the book
directions most minutely.
*Ehrlich method for tubercle bacilli.*--Sections are stained for six
to twenty-four hours in a one per cent. solution of gentian violet,
methyl violet, methyl blue or fuchsine. They will stain more rapidly
if the staining fluid be kept in an incubator at the body temperature.
They should be removed from the staining fluid, and washed in distilled
water, and then transferred (preferably on a glass section lifter) to
a ten per cent. solution of nitric acid in distilled water until they
are nearly decolourised. They should then be very thoroughly washed in
distilled water. They may then be treated with some suitable contrast
stain and mounted in Canada balsam.
*Neelsen’s stain for tubercle bacilli.*--Sections are placed in
Ziehl’s carbol-fuchsine solution (p. 103) which should be warmed for
ten minutes to half-an-hour. They are then decolourised in a solution
of sulphuric acid. Twenty-five per cent. is the strength originally
recommended, but a ten per cent. solution does equally well and injures
the section less. They are then very thoroughly washed in a large
quantity of water, and afterwards may be treated with a contrast stain.
*Gibbes’ double stain for tubercle bacilli.*--
(1) Rosaniline hydrochlorate 2 grms. 25 grs.
Methyl blue 1 grm. 12·5 grs.
Triturate in a glass mortar,
(2) Aniline oil 3 c.c. 37·5 grs.
Rectified spirit 15 c.c. 3-1/2 drms.
Dissolve and add slowly to (1).
(3) Lastly add slowly to the mixture
Distilled water 15 c.c. 3-1/2 drms.
Some of the solution is filtered into a watch glass and warmed. The
sections are placed in it and left for some hours. They are then washed
in methylated spirit till they are sufficiently decolourised, and then
rapidly passed through absolute alcohol and oil of cloves and mounted
in balsam and xylol. It is a very useful stain for examining the sputum
for tubercle bacilli.
In order to stain fluids, such as blood, pus, or sputum, for organisms,
a very thin layer should be obtained by placing a little of the fluid
between two clean cover-glasses and pressing them together. They are
then separated and allowed to dry. The film is fixed by holding the
cover-glass in a pair of forceps, and passing it slowly through the
flame of a spirit lamp two or three times. Films of pus should be
‘cleared’ after fixing by placing them in a twenty per cent. solution
of acetic acid for three minutes.
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