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Epithelial Membrane Antigen (EMA) Antibody Staining Protocol for Immunohistochemistry
Description: Anti-EMA, E29 reacts with a variety of normal epithelial tissues including: secretory and ductal epithelial cells of the skin, breast and other organs of glandular differentiation including sebaceous, esophageal and gastric glands.
Primary Antibody
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Name: EMA Antibody |
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Clone: E29, Mouse anti-Human |
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Supplier: Dako |
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Catalog Number: M0613 |
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Dilution: 1:500 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: Gill's Hematoxylin or Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Cytoplasmic and plasma membrane |
| Images: Search image |
Additional Information:
| Tissue Type: Skin, Breast cancer, Colon cancer |
| Fixation: Formalin fixed paraffin sections, or acetone fixed frozen sections, or cell smears |
| Positive Control: Skin, Breast cancer, Colon cancer |
| 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:
No need to perform antigen retrieval on frozen sections |
References: