Specificity: anti-Mouse IgG (H+L), anti-Rabbit IgG (H+L)
Chromogen/Substrate: 3-Amino-9-ethylcarbazole (AEC)
TL-015-HA: 15mL Hydrogen Peroxide Block (TA-015-HP), 15mL Ultra V Block (TA-015-UB), 15mL Primary Antibody Enhancer (TL-015-PB), 15mL HRP Polymer (TL-015-PH), 15mL AEC Single Solution (TA-015-SA)
UltraVision detection system detects a specific mouse IgG or rabbit IgG antibody bound to an antigen in tissue sections. The specific antibody is located by a universal secondary antibody formulation conjugated to an enzyme-labeled polymer that recognizes mouse and rabbit immunoglobulins. The polymer complex is then visualized with an appropriate substrate/chromogen.
1. Deparaffinize and rehydrate tissue section.
2. Wash 2 times in buffer.
3. If required, incubate tissue in digestive enzyme (or appropriate pretreatment).
4. Wash 4 times in buffer.
5. To reduce nonspecific background staining due to endogenous peroxidase, incubate slide in Hydrogen Peroxide Block for 10-15 minutes.
6. Wash 4 times in buffer.
7. Apply Ultra V Block and incubate for 5 minutes at room temperature to block nonspecific background staining. NOTE: Do not exceed 10 minutes or there may be a reduction in desired stain. (May be omitted if primary antibodies are diluted in buffers containing 5-10% normal goat serum.)
8. Wash (Optional).
9. Apply primary antibody and incubate according to manufacturer's recommended protocol.
10. Wash 4 times in buffer.
11. Apply Primary Antibody Enhancer and incubate for 20 minutes at room temperature.
12. Wash 4 times in buffer.
13. Apply HRP Polymer and incubate for 30 minutes at room temperature. (NOTE: HRP Polymer is light sensitive. Please avoid unnecessary light exposure and store in opaque vial.)
14. Wash 4 times in buffer.
15. Apply ready-to-use AEC Single Solution to tissue. Incubate for 10 minutes. Modify incubation time to optimize staining in your laboratory.
16. Wash 4 times in DI water.
17. Counterstain and cover slip using an aqueous mounting media.
The specificity and sensitivity of antigen detection is dependent on the specific primary antibody used. General References:
1. Shan-Rong Shi, James Guo, Richard J. Cote, Lillian Young, Debra Hawes, Yan Shi, Sandra Thu, and Clive R. Taylor, Applied Immunohistochemistry & Molecular Morphology, Vol 7, 201-208, 1999.
2. Karen Petrosyan, Rosalba Tamayo, and Daisy Joseph, “Sensitivity of a Novel Biotin-free Detection Reagent(PowerVision+) for Immunohistochemistry” J. Histotechnology, Vol 25, 247-250, 2002.