[Histonet] RE: Dab enhancement

From:"Pixley, Sarah \(pixleysk\)"

 Dear Susan:
There are several ways to enhance. We have found the simplest for us is:

Nickel enhancement of DAB:
DAB (0.5 mg/ml) with 0.08% nickel sulfate and 3% H2O2:
Volume needed:		1 ml	

2% nickel sulfate		40 ul	
DAB Solution		1 ml	
30% H2O2			10 ul	
To make: 2% Nickel sulfate, NiSO4.6H2O: 0.9 g into 45 ml E-pure dd H2O = 2% solution. Keep in Fridge.
(Dilute: 1/25 to give a 0.08% solution.)

NOTE: time to getting staining will be much quicker than without nickel. And this solution develops more precipitate than a non-nickel solution, so if you are doing a lengthy DAB staining because you have many samples, you may want to plan to make more DAB solution in the middle of the staining. 

Another easy trick is to treat your sections with cupric sulfate after staining (does not seem to be as dark or enhancing as adding nickel with the DAB--more brown than blue-black).

Post-DAB enhancement
0.4% cupric sulfate (CuSO4·5H2O)
In 0.9% NaCl

Keep cupric sulfate at room temp. in closed glass bottle (good for many years).
After DAB reaction has been completed and rinsed well with PBS or PB, immerse slides for short time, i.e. 5-20 mins, and then rinse with PBS, saline or water. Darkens regular DAB color to blacker shade. 

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