In article <DGJtJu.HJs at psych.toronto.edu> scott at psych.toronto.edu (Brian Scott) writes:
>Subject: Ethanol fixation in BrdU immunohistochemistry?
>I've been looking into bromodeoxyuridine immunohistochemistry for labeling
>neuronal precursors and I've noticed that many of the papers mention using
>ethanol to fix the tissue and not something like paraformaldehyde. I've been
>given a method to follow which uses Lana's fixative which includes
>paraformaldehyde and picric acid (yikes!) and I've been wondering if ethanol
>would do just as well. I've never heard of just using ethanol before. Does
>anyone know if it's ok? Thanks.
Ethanol alone gives lousy structural preservation, except for smears
of cells on slides. It is normally used in mixtures that also contain
acetic acid. It is then very good for nuclear architecture and nucleic
acid immobilization. Formaldehyde (which is what you get from
paraformaldehyde when it dissolves) does not give nuclei of "classical"
appearance, with sharply defined heterochromatin, though it does
leave the DNA and histones in position. Paraffin embedding damages
tissues fixed in any solution of which formaldehyde is the only
ingredient. Frozen sections or resin embedding are OK after formaldehyde
alone. Formaldehyde-picric mixtures give excellent structural
preservation, but the acid mixtures like Bouin's fluid will extract
some nucleic acids (mainly RNA) if allowed to act for a long time (more
than 24 h). The neutral formal-picric mixture of Stefanini, de Martino &
Zamboni is much used for immunohistochemistry, and also preserves
structural detail well in paraffin sections.
Recipes for fixatives can be found in any textbook of
histological technique. It's important to understand the rationale
behind the inclusion of each ingredient in a fixative mixture.
John A. Kiernan
Department of Anatomy
Univ. of Western Ontario
LONDON, Canada N6A 5C1
e-mail: kiernan at uwo.ca
