The objectif of this histological study is the comparison between the different regions of brain in rats. Questions:
- What is the optimal concentration of formaldehyde for the realization of good histological study of the full brain (brain completely fixing) of rats ?
- are there some other reactifes for a good fixation?
- Are there other ways better than the histological study. To study the areas of the brain. Condition of being simple means?
Dear Gasmi! With my pleasure. Please read carefully all three articles (in Attachment).
Dear Gasmi Salim
This depends mainly on the equipments and the environment in your lab.
So I prefer to start with a pilot study using 1-3 rat brain's
1st use 10% neutral buffered formalin.
2nd once the brains are removed from rat skull immediately put on an ice.
3rd sometime intracardiac fixation with gluteraldehyde may help to preserve the specimens content and sometimes fail.
Some researchers prefer cryo fixation for the brains and other this manuver may fail.
After full processing for the pilot specimens examine under your microsocpe to decide which is better for you.
Also, is your next step is immunomarkers or others you will decide.
So pilot study will help you to check the availability at your lab and what is the best for you.
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Dear Gasmi,
During my PhD studies, I have done immunohistochemical and morphological analysis of entire brain, spinal cord and peripheral tissues. I used to to fix the entire animal with fixatives using intracardiac perfusion method, and then further incubate in fixative following dissection. For details please have a look at my thesis listed below.
Best wishes,
Refik Kanjhan
Thesis Molecular physiology of ATP-gated ion channels assembled fro...
@Vladimir A. Kulchitsky : Dear Vladimir, would you mind to eventually correct*) the attachment of the 3 articles (if it is a problem concerning correct initial upload of these articles)....clicking into any of these icons results in an error message saying:
"An error occurred while loading the image. Please try again."
Thanking you in advance, best regards, WM
NB: *): means that you eventually edit your contribution in Answer#01, citing at least the bibliographic reference(s) [ = title Author(s), journal and year of publication....]
Dear Gasmi, to get good staining results in conventianal staining like Nissl's Cresylviolet or immunohistochemistry (IHC) I recommand to do transcardiel perfusion of 4% Paraformaldehyd in PBS pH 7,4. After perfusion 24 postfixation in the same fixativ, PBS (paraffin sectons )or 30% Sacrose for cryoprotetion (froszen sections). With this fixation protocol you stain 3-5µm paraffin sections as well as 30-50 µm free floating frozen sections (IHC). you can use this fixativ evan for immersion fixation. For mice brain this is ok, but for rat brain you have to take care about so called diffusion artefacts. This could happen when tissue blocks are to big for a fast diffusion of the PFA. Good luck!
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