UltraSensitiveTM SAP Immunohistochemistry Staining Procedure
The ready-to-use immunohistochemistry UltraSensitiveTM SAP kit uses a biotin-labeled secondary antibody mixed with streptavidin-conjugated alkaline phosphatase and substrate chromogen to detect antigens in tissues and cells. The staining procedure is as follows:
1. Deparaffinize and hydrate paraffin sections, rinse with tap water;
2. Perform appropriate antigen retrieval on the tissue according to the requirements of the primary antibody;
3. Remove PBS, apply normal non-immune animal serum to the sections, and incubate at room temperature for 10 minutes;
4. Remove serum, apply the primary antibody to the sections, incubate at room temperature for 60 minutes or at 4°C overnight, rinse with PBS three times, 3 minutes each time (3 × 3 minutes);
5. Remove PBS, apply biotin-labeled secondary antibody to the sections, incubate at room temperature for 10 minutes, rinse with PBS 3 × 3 minutes;
6. Remove PBS, apply streptavidin-alkaline phosphatase reagent to the sections, incubate at room temperature for 10 minutes, rinse with PBS 3 × 3 minutes;
7. Remove PBS, apply BCIP/NBT (or AP-Red) chromogenic reagent to the sections for color development;
8. Rinse with distilled water to stop color development, counterstain with nuclear fast red (or hematoxylin), and mount with aqueous mounting medium.