Hi Christine,

We make up our own antibody cocktail of 34ßE12 (DAKO dilution 1:150) and P63
(DAKO 1:400) and stain the section with this cocktail with a Polymer
peroxidise detection system (DAB).(Heat retrieval in a high pH retrieval
solution for 20 min at 100 C.

The stained slides are washed and darkened using a copper sulphate solution.
Placed in a buffer wash and then stained for the second time with P504S
(AMACR/Racemase) at a dilution of 1:150 with a Polymer Alkaline Phosphatase
detection system (Fast Red).
We do this all on the BondMax Immunostainer.
We have been doing this for over 6 months now and did it this way because we
already had all the concentrated antibodies.
Proper fixation is vital.

Hope this helps.

Regards
Barry 
B.Madigan
Pathology Queensland
Royal Brisbane Campus
Australia





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