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Western blot
ImmunohistochemistryAntibody data
- Antibody Data
- Antigen structure
- References [1]
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- Validations
- Western blot [1]
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- Product number
- GTX25797 - Provider product page
- Provider
- GeneTex
- Proper citation
- GeneTex Cat#GTX25797, RRID:AB_380692
- Product name
- PKC gamma (phospho Thr674) antibody
- Antibody type
- Polyclonal
- Reactivity
- Human, Mouse, Rat
- Host
- Rabbit
Submitted references Hint1 knockout results in a compromised activation of protein kinase C gamma in the brain.
Zhang F, Fang Z, Wang JB
Brain research 2015 Oct 5;1622:196-203
Brain research 2015 Oct 5;1622:196-203
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Supportive validation
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- Experimental details
- Extracts of HeLa cells untreated (1-7) or treated with 1?M PMA for 30 minutes (8-14) were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was either left untreated (1-13) or treated with lambda phosphatase (14), blocked with a 5% BSA-TBST buffer for one hour at room temperature, then incubated with the PKCgamma [pT674] antibody for two hours at room temperature in a 1% BSA-TBST buffer, following prior incubation with: the phosphopeptide corresponding to the immunogen from other PKC isoforms (1-9), the phosphopeptide immunogen (10), the non-phosphopeptide corresponding to the phosphopeptide immunogen (11), a generic phosphothreonine-containing peptide (12) or no peptide (13, 14). After washing, the membrane was incubated with goat F(ab)2 anti-rabbit IgG HRP-conjugate and signals were detected. The data show that the phosphopeptide corresponding to PKCgamma [pT674] blocks the antibody signal. The peptides corresponding to PKCalpha [pT657], betaI [pS661] and PKCeta [pS674] also block the antibody signal, suggesting cross-reactivity of the PKCalpha [pT638] antibody with these sites. The antibody signal was not blocked by the peptides corresponding to PKC isoforms betaII [pT660], gamma [pT514], gamma [pT655], delta [pS664], theta [pS695] and zeta [pT560], demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, further verifying that the antibody is phospho-specific.
