[Histonet] IHC staining weaker in...

[Monfils, Paul]

What is your standard fixative?  Have you made any changes in routine fixation recently?  This sounds like a fixation problem.  As a core research facility, we receive all kinds of tissues from various researchers, often in a variety of fixatives. I have noticed the artifact you describe in some tissues fixed in certain slow-penetrating fixatives.  Some aggressive fixatives give very nice preservation in small tissue samples, but when larger tissues are immersed in such fixatives, the rapid fixation of the outer layer of the tissue actually sets up a barrier to penetration of the fixative into the deeper levels. The result is a poorly fixed interior with a very well fixed exterior, and the poorly fixed deeper areas often stain poorly.