BRDU

[Annica Andersson]

Hello,
We have perfused rats with 4%PF/picric acid and now we have problems to get the BrdU AB to work (Harlan, 1:100 for 90 min), 2M HCL 37C for 30 min might not be enough. When develope with DAB the staining looks weak and the ab dont penetrate to the nuklea. Its there a nother way to denature the sectons (12um) we also have big problems with DAB background and that does not īmake it easy to evaluate.
Regards Annica

[richard03]

Citrate buffer antigen retrieval is an alternate method for BrdU antibody

http://www.ihcworld.com/_protocols/epitope_retrieval/citrate_buffer.htm

[dutchmaninvienna]

It is a bit unclear how you have processed the sections, are they cryostate or paraffin sections? If you feel that you have problems with penetration of the antibodies you might 1) increase the incubation times, 2) use detergents, 3) in case of paraffin, increase the deparaffination times. 4) in case of cryostate sections use acetone or chloroform to permeabilize the membranes in your tissues. etc etc. Just shortly write down you protocol and it will be easier to find out your problem.

jan