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CA19-9 (Sialyl Lewis a) Antibody Staining Protocol for Immunohistochemistry
Description: The Lewis blood group antigens Lewisa and Lewisb are carbohydrates that form epitopes on glycolipids and glycoproteins. CA19-9 is an epitope on the sialylated Lewisa carbohydrate structure. Sialylated Lewisa plays a role in cell adhesion by acting as a functional ligand for the inducible adhesion molecule E-selectin.
Primary Antibody
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Name: CA19-9 Antibody |
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Clone: C241:5:1:4, Mouse anti-Human |
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Supplier: Novocastra Labs |
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Catalog Number: NCL-CA19-9 |
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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: 60min/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: 95-100 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: Extracellular membrane-associated |
| Images: Search image |
Additional Information:
| Tissue Type: Small bowel |
| Fixation: Formalin-fixed paraffin sections, or acetone fixed frozen sections |
| Positive Control: Small bowel |
| Negative Control: Omit primary antibody, isotype control, 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:
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There is no need to perform antigen retrieval for acetone fixed frozen sections |
References: