Staining protocol

1. Deparaffinize and rehydrate formalin-fixed paraffin-embedded tissue section.
2. Add enough drops of an appropriate hydrogen peroxide block to cover the sections. Incubate for 10 minutes. Wash 2 times in buffer.
3. Perform appropriate pretreatment if required. Wash 3 times in buffer.
4. Apply an appropriate protein block and incubate for 5 minutes at room temperature to block nonspecific background staining. Wash 1 time in buffer.
5. Apply primary antibody and incubate according to manufacturer's protocol.
6. Wash 4 times in buffer. Apply Biotinylated Goat anti Mouse & Rabbit IgG(H+L) solution (ab64257) and incubate for 10 minutes at room temperature. Wash 4 times in buffer.
7. Apply enzyme labeled streptavidin and incubate according to manufacturer’s protocol.
8. Rinse 4 times in buffer. Place slide in appropriate chromogenic substrate and incubate until desired reaction is achieved. Rinse 4 times in buffer.
9. Add enough drops of Hematoxylin to cover the section. Incubate for 1 minute.
10. Rinse 7-8 times in tap water. Add Mounting Medium to cover the section.

This reagent is intended to be used in a labeled streptavidin-biotin immunoenzymatic antigen detection system. This technique involves the sequential incubation of the specimen with an unconjugated primary antibody specific to the target antigen, a biotinylated secondary antibody which reacts with the primary antibody, enzyme-labeled streptavidin, and substrate-chromogen.