IHC Detection Kit for Mouse and Rabbit IgG Primary Antibody

• HRP-conjugated Secondary Antibody: 50 ml
• 5% BSA Blocking Buffer: 50 ml
• 3% Hydrogen Peroxide: 50 ml

• PBS for IHC
• Chromogenic Solution
• Hematoxylin
• Mounting Medium

Protocol for paraffin sections

1. Dewax paraffin sections to water using a standard protocol.
2. Incubate with 3% Hydrogen Peroxide in deionized water at room temperature for 5-10 minutes to eliminate endogenous peroxidase activity. Rinse with PBS for 5 minutes, repeat for a total of 3 rinses.
3. Perform antigen retrieval according to requirements, heat retreival or enzyme retrieval can be used. This step can also be omitted completely.
4. Block the sample using 5% BSA blocking buffer, incubate at 37°C for 30 mins. Gently shake the sample to dry, do not wash.
5. Add the primary antibody (Rabbit/Mouse IgG), incubate at 37°C for 1-2 hours or overnight at 4°C. Rinse with PBS for 5 minutes, repeat for a total of 3 rinses.
6. Add the HRP-conjugated secondary antibody, incubate at 37°C for 30 minutes. Rinse with PBS for 5 minutes, repeat for a total of 3 rinses.
7. Develop with chromogenic solution (DAB or AEC), using a microscope to observe and control reaction time according to requirements. Rinse thoroughly when the required intensity is achieved.
8. Re-dye with hematoxylin for 0.5-2 minutes as required. Dehydrate and clear the sample.
9. Mount the slide with natural resin, water-soluble mounting medium or mounting medium.

• For heat retrieval, citrate buffer, EDTA solution or PBS can be used as antigen retrieval solutions.
• For enzyme retrieval, composite retrieval solution, antigen retrieval solution (used in conjunction with heat retrieval), trypsin, and pepsin can be used to digest tissue.