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Fas Antibody Staining Protocol for Immunohistochemistry
Description:
Cytotoxic T lymphocyte
(CTL)-mediated cytotoxicity constitutes an important component of
specific effector mechanisms in immunosurveillance against
virus-infected or -transformed cells. Two mechanisms appear to
account for this activity, one of which is the perforin-based
process. Independently, a FAS-based mechanism involves the
transducing molecule FAS (APO-1) and its ligand (FAS-L). The human
FAS (APO-1) protein is a 48 kDa cell surface glycoprotein that
belongs to a family of receptors that includes CD40, nerve growth
factor receptors and tumor necrosis factor receptors. The FAS
antigen is expressed on a broad range of lymphoid cell lines,
certain of which undergo apoptosis in response to treatment with
antibody to FAS. These findings strongly imply that targeted cell
death is potentially mediated by the intercellular interactions of
FAS with its ligand or effectors, and may be critically involved in
CTL-mediated cytotoxicity.
Primary Antibody
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Name: Fas Antibody |
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Clone: B-10, Mouse anti-Human |
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Supplier: Santa Cruz Biotechnology |
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Catalog Number: sc-8009 |
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Dilution: 1:100 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: Cytoplasmic |
| Images: Search image |
Additional Information:
| Tissue Type: Testis, colon, spleen. |
| Fixation: Formalin fixed paraffin sections. |
| Positive Control: Testis, colon, spleen. |
| 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: