Re: Mulligan's
Dolors Fuster wrote:
>
> Does anybody have a protocol for Mulligan's stain?
> I can't find it on the net
That's not surprising, but henceforth it will be there.
In the following instructions, "water" means tap water.
1. Cut slices of formaldehyde-fixed (at least 2-3 weeks)
brain.
2. Refix in 10% formalin, 8 hours
3. Wash, running water, 12 hours
4. Put in a solution of phenol (4%), copper sulphate (0.5%)
and conc HCl (0.125%), heated to 60°C, for 2 minutes.
(This makes a film over white matter that protects it
from the next reagent.)
5. Wash in a large volume of cold water, 1 minute.
6. Put in 2% aqueous tannic acid, 1 minute.
(This binds to proteins in grey matter.)
7. Wash, running water, 5 minutes.
8. Put in 2% iron alum (ferric ammonium sulphate)
until the grey matter is black (usually about
a minute).
9. Wash in running water for 24 hours.
The original paper is well worth reading, because it
includes more detailed explanations and instructions,
and also describes a way of mounting the stained slices.
Mulligan, J.J. 1931. A method of staining the brain for
macroscopic study. Journal of Anatomy 65: 468-473.
An alternative technique that you might consider is
interrupting the method after Step 5 and then
immersing in a 1% aqueous solution of copper(II)
phthalocyaninetetrasulphonic acid tetrasodium salt
(Aldrich 27,401-1) for 10-15 minutes. Wash in running
water for at least 15 minutes but not longer than
48 hours. Store the stained slices in water with
0.5% acetic acid. They can be kept for at least
4 years, and probably much longer. The blue dye,
despite its long name, costs only $30 for 50 grams.
The 1% solution, which is opaque (looks like blue
paint) keeps well for at least 2 years and can be
used repeatedly. A paper by Michael Wu and me,
describing this method, is in the press
(Biotechnic & Histochemistry) and should be
in print before the end of the year.
-------------------------
John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London, Canada N6A 5C1
kiernan@uwo.ca
http://publish.uwo.ca/~jkiernan/
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