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Estrogen Receptor (ER) Antibody Staining Protocol for Immunohistochemistry
Description: Recognizes a protein of 67kDa, which is identified as estrogen receptor (ER) alpha. The ER gene consists of more than 140kb of genomic DNA divided into 8 exons, being translated into a protein with six functionally discrete domains, labeled A through F. This antibody strongly stains the nucleus of epithelial cells in breast carcinomas. The ER is an important regulator of growth and differentiation in the mammary gland. Presence of ER in breast tumors indicates an increased likelihood of response to anti-estrogen (e.g. tamoxifen) therapy.
Primary Antibody
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Name: Estrogen Receptor (ER) Antibody |
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Clone: SP1, Rabbit anti-Human |
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Supplier: Lab Vision |
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Catalog Number: RM-9101-S |
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Dilution: 1:200 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed. |
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Incubation Time/Temp: 60 minutes/room temperature |
Antigen Retrieval
| Device: IHC-TekTM Epitope Retrieval Steamer Set (Cat# IW-1102) |
| Buffer/pH value: IHC-TekTM Epitope Retrieval Solution (Cat# IW-1100) |
| Heat/Cool Temperature: 95 ºC - 100 ºC/room temperature |
| Heat/Cool Time: 20 minutes/20 minutes |
Detection Methods
| Standard Method: ABC Method or LSAB Method |
| Enhanced Method: Polymeric Methods |
Chromogen Substrate
| Reagent: DAB |
| Incubation Time/Temperature: 1-3 minutes/room temperature |
Counterstain
| Reagent: Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Nuclear |
| Images: Search image |
Additional Information:
| Tissue Type: Breast carcinomas, MCF-7 cells. |
| Fixation: Formalin fixed paraffin sections |
| Positive Control: Breast carcinomas, MCF-7 cells. |
| Negative Control: Omit primary antibody, isotype control or absorption control |
| Blocking: 2-5% normal serum to reduce unspecific background staining; 0.5-3% H2O2 to block endogenous peroxidase activity; avidin/biotin to block endogenous biotin activity if necessary |
Notes:
Not tested on frozen sections |
References:
1. Katoh AK, Stemmler N, Specht S, D'Amico F (1997) Immunoperoxidase staining for estrogen and progesterone receptors in archival formalin fixed, paraffin embedded breast carcinomas after microwave antigen retrieval. Biotech Histochem. 72(6):291-8. PubMed Abstract