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Phospho-p44/42 Map Kinase Antibody Staining Protocol for Immunohistochemistry
Description:
Both p44 and p42 MAP
kinases (Erk1 and Erk2) function in a protein kinase cascade that
plays a critical role in the regulation of cell growth and
differentiation. MAP kinases are activated by a wide variety of
extracellular signals including growth and neurotrophic factors,
cytokines, hormones and neurotransmitters. Activation of MAP kinases
occurs through
phosphorylation of threonine and tyrosine (202 and 204 of human MAP
kinase [Erk1] or 183 and 185 of rat Erk2) at the sequence T*EY* by a
single upstream MAP kinase kinase (MEK) (5,6). Both kinases are
known to weakly autophosphorylate on tyrosine.
Primary Antibody
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Name: Phospho-p44/42 Map Kinase (Thr202/Tyr204) Antibody |
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Clone: Rabbit polyclonal |
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Supplier: Cell Signaling Technology |
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Catalog Number: 9101 |
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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, hamster, chicken, zebra fish |
| Fixation: Formalin fixed paraffin sections, or acetone fixed frozen sections (no antigen retrieval is needed) |
| Positive Control: Ischemia brain |
| 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: