Hello, I am using cell signaling technology p-Akt IHC kit(catalog number8100). The kit uses rabbit primary antibody against p-AKt Ser 473,biotinylated secondary antibody with avidin DH & HRP & Nova Red as a substrate.
My problem:1) Using the kit Non specific nuclear staining of strong intensity in normal tissues . This staining persists to a mild degree in negative control slides(where I omit primary antibody). AS a counter stain we are dipping the slides in Gills hematoxylin for 2 dips(should we increase this)
2) I am unable to use their primary pre diluted antibody with DAKO envison kit so as to get final staining with DAB.I tried using all the kit components till the nova red substrate chromogen step where I substituted DAB but it did not work. Then I tried using their pre diluted antibody with DAKO envison kit :!: :?:
I need urgent help on this. Anyone got any suggestions. If anyone has used the CST kit on FFPE sections their experience will be invaluable.[/b]