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
*Schäfer’s modification* of Pal’s method.--In this method hardening in
Müller’s fluid for three or four weeks is sufficient. The sections are
made exactly as in the previous method, and transferred to Marchi’s
fluid (p. 24) for six hours. They are washed and stained all night in
the following:--
Hæmatoxylin 1 4 grs.
Alcohol 10 45 min.
Acetic acid (2 per cent. aqueous solution) 100 1 oz.
The subsequent processes of differentiation, bleaching, &c., are
exactly the same as in Pal’s method.
*Osmic acid.*--Employed with fresh and also with hardened specimens to
demonstrate the medullary sheath. Much the best results are obtained
with the former. The nerves, or small pieces of the central nervous
system are placed in half to one per cent. solution of osmic acid as
soon as possible after death and kept in the dark for about a week.
The tissue must be very thoroughly washed in running water to remove
all traces of osmic acid, and then stained for a couple of days in
borax carmine to demonstrate the nuclei and axis cylinders. Sections
may be made in gum, or the tissue may be teased with needles and then
mounted in Farrant. Embedding in celloidin, and mounting in balsam are
inadvisable, because the ether tends to dissolve out myelin, and the
clarifying oil to render it too transparent.
2. *Intra-muscular ramifications of nerves*:--
*Sihler’s chloral hæmatoxyline method.*--This method reveals the
intra-muscular nerve-endings, and also brings into prominence the
curious “muscle spindles” which Sherrington has shown to be connected
with the posterior nerve roots, and which are believed by some to be
the end organs subserving muscular sense.
A piece of muscle is taken as soon as possible after death, or from an
amputated limb, and slices cut about one-tenth of an inch thick with
the freezing microtome. Transfer for twenty-four hours to the following
solution:--
Acetic acid 1 part.
Glycerine 1 "
One per cent. aqueous solution of chloral hydrate 6 parts.
The tissues swell up in this fluid and become translucent and
gelatinous in appearance. They are now placed in pure glycerine until
saturated as shown by their sinking to the bottom of the dish. This
usually takes several days. They may now be stained in the following
solution:--
Ehrlich’s hæmatoxyline (p. 70) 1 part.
Glycerine 1 "
One per cent. aqueous solution of chloral hydrate 6 parts.
They may be left in this from three days to a week with little fear of
overstaining. Portions may then be teased with needles, and mounted in
glycerine, or the stained tissue may be pressed out into a sufficiently
thin layer by squeezing it forcibly between two glass slides.
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