By "fresh" do you mean not chemically fixed, just frozen ?

Most antigen retrieval procedures break the chemical bonds between the chemical fixative and the tissue fixed This allows access to the antigens for the antibodies. If your material is not chemically fixed, why do you need antigen retrieval ? Is the material you use for freezing (OCT or ?) interfering with your labeling ? If this is the issue, try using a longer wash step prior to incubation with the primary.

Hi

Thanks for your answer. I guess my question was more about the possibility to use this method (after a quick PFA fixation). I have the feeling that most people use the antigen retrieval on FFPE samples, no?

Depends on your antigen and antibody. I have a number of antibodies that work very well without antigen retrieval on PFA andPFA/ glutaraldehyde fixed material. Try a run without antigen retrieval and see what happens.

I normally use the microwave antigen retrieval method in an old normal household microwave. I have also used sodium metaperiodate saturated in water; sections exposed to it for 10 minutes and then sections washed a few times for a couple of minutes prior to antibody incubation.

Great! Thanks I will give it a try!