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TGase1 (Transglutaminase 1) Antibody Staining Protocol for Immunohistochemistry
Description: Terminally differentiating mammalian epidermal cells acquire an insoluble, 10 to 20 nm thick protein deposit on the intracellular surface of the plasma membrane known as the cross-linked cell envelope (CE). The CE is a component of the epidermis that is generated through formation of disulfide bonds and gamma-glutamyl-lysine isodipeptide bonds, which are formed by the action of transglutaminases (TGases). TGases are intercellularly localizing, Ca2+-dependent enzymes which catalyze the formation of isopeptide bonds by transferring an amine onto glutaminyl residues, thereby cross-linking glutamine residues and Lysine residues in substrate proteins. TGases influence numerous biological processes including blood coagulation, epidermal differentiation, seminal fluid coagulation, fertilization, cell differentiation and apoptosis.
Primary Antibody
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Name: TGase1 (N-20) |
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Clone: Polyclonal Goat ant-Human |
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Supplier: Santa Cruz Biotechnology |
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Catalog Number: sc-18127 |
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Dilution: 1:50 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 ºC - 100 ºC/room temperature |
| Heat/Cool Time: 20 minutes/20 minuters |
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: Intercellular |
| Images: Search image |
Additional Information:
| Species Reactivity: Human, mouse, rat |
| Fixation: Formalin fixed paraffin sections. |
| Positive Control: Skin (epidermis) |
| 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: