GTX10564

antibody from GeneTex

Targeting: CREB1

Immunocytochemistry

Chromatin Immunoprecipitation

Antibody data

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Validation data

CREB1 protein structure - GTX10564 shown in red.

Supportive validation

Submitted by: GeneTex (provider)
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Experimental details: Whole cell extracts of human A549 cells (approximately 20,000 cells per lane) untreated (1), stimulated with EGF (2) or treated with pervanadate (3), were resolved by SDS-PAGE and transferred to PVDF.?Whole cell extracts of human A549 cells (approximately 20,000 cells per lane) untreated (1), stimulated with EGF (2) or treated with pervanadate (3), were resolved by SDS-PAGE and transferred to PVDF. The membrane was blocked with a casein/Tween 20 buffer, then incubated with mouse anti-CREB/ATF1 [pS133] antibody (GTX10564) at 0.5 ?g/mL for 1 hour at room temperature. After washing, the membrane was incubated with an anti-mouse HRP-conjugated secondary antibody and signals were detected using an ECL detection method (exposure time: 30 seconds).

Supportive validation

Submitted by: GeneTex (provider)
Main image
Experimental details: "Up-regulation and Antibody-Peptide Competition.? Peptide Competition and Phosphatase Treatment. Extracts of NIH3T3 cells untreated (1) or treated with 50ng/mLPDGF for 15 minutes (2-6) were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer for one hour at room temperature and either left untreated (1-5) or treated with lambda phosphatase (6), then incubated with the CREB (GTX10564) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 2, 6), the non-phosphopeptide corresponding to the phosphopeptide immunogen (3), a generic phosphoserine-containing peptide (4), or the phosphopeptide immunogen (5). After washing, the membrane was incubated with goat F(ab¡¦)2 anti-rabbit IgG HRP conjugate and signals were detected. The data show that only the phosphopeptide corresponding to CREB [pSpS129/133] blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, further verifying that the antibody is phospho-specific. Note: Pretreatment of cells with PKA inhibitor (PKI) or GSK-3beta inhibitor (LiCl) diminished the signal, indicating the dual phosphoreactivity of this antibody (data not shown)."