I agree with Carl Hobbs: "Perhaps your H2O2 ( substrate) stock has 'gone off'?
That will give you a weaker/negative DAB result."
If your frozen DAB aliquots keep the same color, I would be very suspect of the
Hydrogen Peroxide. It can become unstable very quickly.
Terri L. Braud, HT(ASCP)
HNL
I do manual IHC-DAB regularly
DAB shouldn't "go off" after freezing unless you have somehow oxidised it
during mixing/aliquoting.
It would then show as a dark brown aliquot
I state this because I still make up my own DAB from dry powder ( have been
doing for > 20yrs)
SIgma D5637
I dissolve,
