Hello, I intend to prepare rat's fetal and pup brains for immunohistochemistry and immuofluorescence. I am required to inject the dams with BrDu, but I don't know how to prepare the solution. Can somebody help out with the protocol? Secondly, since I am travelling with the tissues to another lab, I am required to preserved them in 4% PFA and after fixation in 0.1% sodium axide. How do I prepare the sodium axide, and for how long can the tissues be stored in it before it goes bad? Please I need your assistance.
Brdu solubility in saline is 10 mg per ml. See data sheet:
https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Product_Information_Sheet/b5002pis.pdf
It should be fairly easy to inject the Brdu in saline IP in rats. You can use a fairly large volume if you need to, as much as 5-15 ml depending on the size of the rat.
As to the fixation, the PFA should of course be buffered. Not sure how long you should do the PFA fixation. That would depend on the method that you use, i.e. immersion or perfusion.
Also, I am not familiar with use of sodium azide following PFA fixation, but it may be common to do this in such experiments. Sodium azide can be made up in water or saline at a final concentration of 0.1% at which concentration it is highly effective as an anti-microbial. Just be very careful when you prepare, use, and transport the sodium azide solution as it is a very potent metabolic poison. Use gloves to handle it and promptly clean up any spills. Azide is highly reactive with certain metals like copper and lead. You should check to make sure that there is no interaction between PFA and azide.
Also, azide is very stable in aqueous solution. It should remain good for many months.
Thank you Robert for these information. I will follow your advise strictly
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