- Antigen retrieval buffer
- Hydrogen peroxidase block
- High effect immunoblock
- Mouse probe HRP
- DAB chromogen (20X DAB buffer)
- Counter strain Hematoxylin
- Antibody dilution buffer
|KIT CONTENTS||Description||Format||Recommend time|
|Hydrogen Peroxide Block||3% hydrogen peroxide solution with less than 0.1% Ready to Use sodium azide.||Ready to Use||10-20 minutes.|
|Immunoblock||PBS solution, pH 7.6, with 0.5% BSA, and less than 0.1% Ready to Use sodium azide.||Ready to Use||10-30 minutes.|
|Mouse Probe HRP||Conjugated HRP Reactive with mouse primary Ready to Use antibodies.||Ready to Use||30-45 minutes.|
|DAB Chromogen (20x)||3,3' Diaminobenzidine (DAB) chromogen||Concentrate||Diluted with DAB Buffer (chromogen 1part: buffer 19 parts)|
|DAB Buffer||Substrate buffer, pH 7.5 with hydrogen peroxide||Ready to Use||1-10 minutes.|
|Hematoxylin||Hydrogen peroxide||Ready to Use||0.5-1 minutes.|
|Antigen retrieval buffer||20X Citrate Buffer (pH6)||20X||
|Antibody dilution buffer||Primary antibody and incubate according to manufacturer's protocol||Ready to Use||
Staining Protocol :
1. Deparaffinize and rehydrate formalin-fixed paraffin-embedded tissue section.
2. Add enough drops of Hydrogen Peroxide Block to cover the sections. Incubate for 10 minutes. Wash 2 times in PBS buffer.
3. Perform appropriate pretreatment if required. Wash 3 times in PBS buffer.
4. Apply Immunoblock and incubate for 10-30 minutes at room temperature to block non-specific background staining. Wash 2 times in PBS buffer.
5. Apply mouse primary antibody and incubate according to manufacturer's protocol.
6. Wash 3 times in PBS buffer.
7. Apply Mouse Probe HRP and incubate for 30 minutes at room temperature. Wash 3 times in PBS buffer.
8. Add 50 μl DAB Chromogen to 950ul of DAB Buffer (20X dilution), mix by swirling and apply to tissue. Incubate for 1-10 minutes. Rinse 4 times in PBS buffer.
9. Rinse slide in tap water.
10. Apply Hematoxylin counterstain according to manufacturer’s instructions (optional).
11. Dehydrate if required and cover slips.