Thanks so much to everyone who responded! A couple people have suggested that this issue may be due to a disruption to the charge on the slide (due to high temp baking, water bath additives, etc) which causes a hydrophobic barrier to form and prevents reagent from reaching the tissue. This sounds very convincing to me, but we have found that repeated antigen retrieval using a pressure cooker (our standard retrieval for this particular primary) has allowed 2 of the 3 slides to stain with both DAB and Hematoxylin. Would repeated retrieval be able to reverse the slide charge back to normal?
I'm having an issue with some recent samples and was wondering if anyone else has experienced the same thing...I stained some samples with a previously optimized protocol for anti-human Granzyme B on the Ventana Benchmark and about 1/3 of my samples did not stain at all with either GranzB or Hematoxylin. My controls and samples that did stain positive for GranzB look fine and the machine has been recently serviced and working well with other runs. I'd really appreciate your input if you have any advice or suggestions on what could have gone wrong. Thanks, Anna _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet