EliVisionTM Super/HRP Immunohistochemistry Staining Procedure
The EliVisionTM super/HRP kit is a further development of the EliVisionTM plus/HRP kit. It is based on polymer technology, which conjugates multiple anti-mouse or/and anti-rabbit IgG molecules with horseradish peroxidase onto a polymer. This polymer binds to the primary antibody, thereby amplifying the signal of antigen-antibody binding. The staining procedure is as follows:
1. Deparaffinize and hydrate paraffin sections, then rinse with tap water;
2. Perform appropriate antigen retrieval on the tissue according to the requirements of the primary antibody;
3. If necessary (if the tissue contains endogenous peroxidase), apply peroxidase blocking reagent to the sections, incubate at room temperature for 10 minutes, and rinse with PBS three times, each for 3 minutes (3 × 3 minutes);
4. Remove PBS, apply the primary antibody to the sections, incubate at room temperature for 60 minutes or at 4°C overnight. Rinse with PBS 3 × 3 minutes;
5. Remove PBS, apply the amplifier to the sections, incubate at room temperature for 10 minutes, and rinse with PBS 3 × 3 minutes;
6. Remove PBS, apply the polymer enzyme conjugate to the sections, incubate at room temperature for 10 minutes, and rinse with PBS 3 × 3 minutes;
7. Remove PBS, apply freshly prepared DAB or AEC chromogenic reagent to the sections for color development;
8. Rinse with tap water to stop color development, counterstain with hematoxylin (if necessary, differentiate with 1% hydrochloric acid ethanol), and return to blue with PBS. If DAB is used for color development, dehydrate the sections through a graded ethanol series, clear with xylene, and mount with neutral balsam. If AEC is used for color development, the sections cannot be dehydrated with ethanol and should be directly mounted with an aqueous mounting medium.