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VEGF Antibody Staining Protocol for Immunohistochemistry
Description: The onset of angiogenesis is believed to be an early event in tumorigenesis and may facilitate tumor progression and metastasis. Several growth factors with angiogenic activity have been described. These include fibroblast growth factors (FGFs), platelet derived growth factor (PDGF) and vascular endothelial growth factor (VEGF). VEGF is a dimeric glycoprotein with structural homology to PDGF. Several variants of VEGF have been described that arise by alternative mRNA splicing. It has been speculated that VEGF may function as a tumor angiogenesis factor in vivo because the expression pattern of VEGF is consistent with a role in embryonic angiogenesis. VEGF mRNA is formed in some primary tumors, VEGF is produced by tumor cell lines in vitro and VEGF mitogenic activity appears to be restricted to endothelial cells. A member of the PDGF receptor family, Flt, has been identified as a highaffinity receptor for VEGF.
Primary Antibody
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Name: VEGF (147) Antibody |
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Clone: Rabbit polyclonal |
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Supplier: Santa Cruz Biotechnology |
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Catalog Number: sc-507 |
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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 min/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) or IHC-TekTM Proteinase K Solution (Cat# IW-1101) |
| 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: Cytoplasmic/membrane |
| Images: Search image |
Additional Information:
| Species Reactivity: Human, mouse, rat |
| Fixation: Formalin fixed paraffin sections |
| Positive Control: Tumor cells in hemangiosarcomas |
| 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 |
References: