trying to stain m1 microglia in AD human brain tissue, formalin fixed and parrafin embedded 10 years ago (pretty old)

-jc Virus as positiv control

-trs ph 9,0 pressure cooker 5 min

-dilution buffer for antibody: bsa 1 % tbs ph 7,6  

-antibody 1:300 dilution.

using this protocol my positve control got good results,

but not the AD

lots of Background staining,

reactive Neurons and Astro´s are also slightly positive

and almost none positve microglia (braak 4-5)

what could i Change to get a more specific result?

raising the antibody Dilution to 1:200 even though its the best Dilution for my positive conrtol?

raising the bsa to 5% to get rid of the Background?

thanks in advance

Tim Hartmann