[Histonet] DAB Chromogen

[Nancy Herman]

I have this DAB staining protocol for IHC.  It was used on the DAKO autostainer and incubated for 5min.  Our controls using the DAKO DAB chromogen kit worked fine.  Does anyone have any experience with a DAB protocol similar to this or have any other information to add to this protocol?  Is incubating on ice necessary?

TRIS-DAB:

200 ml TRIS-DAB, 0.05M, Ph 7.6
40 mg DAB (aliquots in 1.5 ml, -20 Celsius)

filter
incubate slides (eventually on ice)
add: 34 ul H2O2 (30%)

stop reaction in PBS
check slides under the microscope

Thanks,
Nancy
Animal Disease Research Laboratory
Canada