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TGF beta RI Antibody Staining Protocol for Immunohistochemistry
Description: A total of three members of the TGF ß family, TGFß1, TGFß2 and TGFß3, have been identified in mammals. Each is synthesized as a latent precursor that is subsequently cleaved forming the 112 amino acid growth factor which becomes active upon dimerization. TGFßs mediate their activity by high affinity binding to the type II receptor 70 kDa transmembrane protein with a cytoplasmic serine-threonine kinase domain. For signaling growth inhibition and early gene responses the type II receptor requires both its kinase activity and association with a 53 kDa TGFß-binding protein, designated the type I receptor. Two independent groups have recently described the cloning and sequence analysis of genes encoding 53 kDa TGFß type I receptor proteins (94 kDa receptor complex) designated ALK-5 (TßR-1) and TSR-1, respectively.
Primary Antibody
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Name: TGF beta RI (V-22) Antibody |
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Clone: Rabbit polyclonal |
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Supplier: Santa Cruz Biotechnology |
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Catalog Number: sc-398 |
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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 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) |
| 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 |
| Images: Search image |
Additional Information:
| Species Reactivity: Human, mouse, rat |
| Fixation: Formalin fixed paraffin sections |
| Positive Control: Prostate |
| 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: