Protocol for AEC Peroxidase Substrate Solution




AEC (3-amino-9-ethylcarbazole) Peroxidase Substrate Solution Red


Final Dilution:


       0.05% AEC and 0.015% H2O2 in 0.05 M Acetate Buffer, pH 5.5


Stock Solutions:


       1% AEC (20x) in 100% DMF (N, N Dimethylformamide):

       Dissolve 0.1 g of AEC in 10 ml of 100% DMF and store this solution at 4 °C.


       0.3% H2O2 (20x) in Distilled Water:

       Add 100 ul of 30% H2O2 in 10 ml distilled water and store at 4 °C.


       1M Acetate Buffer (20x), pH 5.5:

 Sodium acetate, trihydrate (MW 136.1) ------ 13.6 g

 Distilled water ----------------------------------- 100 ml

 Mix to dissolve and adjust pH to 5.5 using concentrated HCl or glacial acetic acid.


       0.05M Acetate Buffer (1x), pH 5.5:

       1M Acetate buffer ------------------------------- 1 ml

       Distilled water ----------------------------------- 19 ml

       Mix well and adjust pH if necessoary.


Working Solution:


       Add 5 drops of 1% AEC to 5 ml of 0.05M Acetate Buffer, pH5.5 and mix well.

       Add 5 drops of 0.3% H2O2 and mix well.

       Incubate sections for 10-15 minutes at room temperature.


Note: AEC color products can be washed off with ethanol/alcohol solution. So slides must be coverslipped using aqueous mounting media.