[Histonet] BrdU Indirect IF staining

[Julia Edgar]

Dear Teri

Here's a protocol I have used in the past. However, I don't know if it inhibits DAPI staining (but see below). For cryostat sections of tissue perfused with paraformaldehyde or similar, stain your sections with the other antibody (primary and secondary) then fix sections in 50% acetic acid/50% ethanol for 10 minutes and wash thoroughly in PBS. Treat sections with 50% HCl/1% Triton-X 100 for 10 minutes, room temperature to denature the DNA. Wash thoroughly in PBS then bock and apply the anti-BrdU antibody for 2 hours, room temperature, in blocking solution with 0.2% Triton-X 100. Wash and apply secondary antibody.

On cells, I have used 0.07M NaOH (made fresh) to denature the DNA. This does NOT inhibit DAPI staining, but I have not used this method on tissue sections.

Julia

Julia Edgar BSc (Hons), PhD
University of Glasgow Veterinary School