Recombinant Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR933Y] to GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279)
- Suitable for: WB, IP, IHC-P, Dot blot
- Reacts with: Mouse, Rat, Human, Zebrafish
Related conjugates and formulations
Overview
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Product name
Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] -
Description
Rabbit monoclonal [EPR933Y] to GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) -
Host species
Rabbit -
Specificity
This antibody detects both GSK alpha phosphorylated on tyrosine 279 and GSK3 beta phosphorylated on tyrosine 216.
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Tested applications
Suitable for: WB, IP, IHC-P, Dot blotmore details
Unsuitable for: Flow Cyt or ICC/IF -
Species reactivity
Reacts with: Mouse, Rat, Human, Zebrafish -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HEK-293 cell lysate. Cell lysate from untreated, nerve growth factor-beta treated PC-12 and SH-SY5Y cells, Zebrafish lysates and mouse brain lysates. IHC: Human brain tissue, glioma, thyroid carcinoma and ovarian carcinoma.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 40% Glycerol (glycerin, glycerine), 0.05% BSA, 59% PBS -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR933Y -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Isotype control
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab68476 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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WB |
1/500 - 1/2000. Detects a band of approximately 47-52 kDa (predicted molecular weight: 47-52 kDa).
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IP |
1/40.
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IHC-P |
1/50. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
This antibody may not be suitable for IHC with mouse, rat or zebrafish samples. See IHC antigen retrieval protocols. For unpurified use at 1/100 - 1/250. |
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Dot blot |
Use at an assay dependent concentration.
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Notes |
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WB
1/500 - 1/2000. Detects a band of approximately 47-52 kDa (predicted molecular weight: 47-52 kDa). |
IP
1/40. |
IHC-P
1/50. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. This antibody may not be suitable for IHC with mouse, rat or zebrafish samples. See IHC antigen retrieval protocols. For unpurified use at 1/100 - 1/250. |
Dot blot
Use at an assay dependent concentration. |
Target
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Cellular localization
GSK3 beta: Cytoplasm. Nucleus. Cell membrane. The phosphorylated form shows localization to cytoplasm and cell membrane. The MEMO1-RHOA-DIAPH1 signaling pathway controls localization of the phosophorylated form to the cell membrane. -
Database links
- Entrez Gene: 2931 Human
- Entrez Gene: 2932 Human
- Entrez Gene: 56637 Mouse
- Entrez Gene: 606496 Mouse
- Entrez Gene: 50686 Rat
- Entrez Gene: 84027 Rat
- Entrez Gene: 30654 Zebrafish
- Entrez Gene: 30664 Zebrafish
see all
Images
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid carcinoma tissue labelling GSK3 (alpha + beta) with purified ab68476 at 1/2000. Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes. ab209101, a Rabbit specific IHC polymer detection kit HRP/DAB was used. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
Positive staining on human thyroid carcinoma without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B).
The section was incubated with ab68476 for 30 minutes at room temperature. The immunostaining staining was performed on a Leica Biosystems BOND® RX instrument.
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All lanes : Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476) at 1/1000 dilution (purified)
Lane 1 : Zebrafish lysates
Then the membrane was incubated with phosphatase
Lane 2 : Untreated Zebrafish lysates
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 47-52 kDa
Observed band size: 47-52 kDaBlocking and diluting buffer: 5% NFDM/TBST
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human ovarian carcinoma tissue labelling GSK3 (alpha + beta) with purified ab68476 at 1/2000. Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes. ab209101, a Rabbit specific IHC polymer detection kit HRP/DAB was used. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
Positive staining on human ovarian carcinoma without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B).
The section was incubated with ab68476 for 30 minutes at room temperature. The immunostaining staining was performed on a Leica Biosystems BOND® RX instrument.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human glioma tissue sections labeling GSK3 (alpha + beta) with Purified ab68476 at 1:50 dilution (5.3 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
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All lanes : Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476) at 1/1000 dilution (purified)
Lane 1 : Mouse brain lysates.
Then the membrane was incubated with phosphatase
Lane 2 : Untreated Mouse brain lysates
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 47-52 kDaBlocking and diluting buffer: 5% NFDM/TBST
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All lanes : Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476) at 1/500 dilution (unpurified)
Lane 1 : SH-SY5Y (Human neuroblastoma epithelial cell) whole cell lysates
Lane 2 : SH-SY5Y (Human neuroblastoma epithelial cell) treated with nerve growth factor at 100ng/ml for 5 minutes. Whole cell lysates.
Lane 3 : SH-SY5Y (Human neuroblastoma epithelial cell) treated with nerve growth factor at 100ng/ml for 5 minutes. Whole cell lysates. Then the membrane was incubated with phosphatase.
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 47-52 kDa
Observed band size: 47-52 kDa
Exposure time: 10 secondsBlocking and diluting buffer and concentration: 5% NFDM/TBST.
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Human brain tissue stained with unpurified ab68476 at 1/100 dilution.
Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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GSK3 (alpha + beta) (phospho Y216 + Y279) was immunoprecipitated from 10μg HeLa (human cervix adenocarcinoma) whole cell lysate with ab68476 at 1/50 dilution (2μg in 0.35mg lysates). Western blot was performed from the immunoprecipitate using ab68476 at 1/200 dilution (9 μg/ml). VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.
Lane 1 (Input): HeLa (human cervix adenocarcinoma) treated with 1uM staurosporine for 4h whole cell lysate 10μg
Lane 2 (+): HeLa (human cervix adenocarcinoma) treated with 1uM staurosporine for 4h whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab68476 in HeLa (human cervix adenocarcinoma) treated with 1uM staurosporine for 4h whole cell lysateExposure Time: 30 seconds
Blocking and diluting buffer and concentration: 5% NFDM/TBST. -
All lanes : Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476) (unpurified)
Lane 1 : PC12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate
Lane 2 : Whole cell lysate from PC12 (Rat adrenal gland pheochromocytoma cell line) cells treated with nerve growth factor-ß at 100ng/ml for 5 minutes.
Lane 3 : Whole cell lysate from PC12 (Rat adrenal gland pheochromocytoma cell line) cells ) treated with nerve growth factor-ß at 100ng/ml for 5 minutes. Membrane incubated with phosphatase
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG H+L (HRP))
Developed using the ECL technique.
Predicted band size: 47-52 kDa
Exposure time: 3 secondsBlocking and dilution buffer: 5% NFDM/TBST
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Dot Blot analysis of Lane 1: Human GSK3 (alpha + beta) (pY216 + pY279) phospho peptide and Lane 2: Human GSK3 (alpha + beta) non-phospho peptide labeling GSK3 (alpha + beta) (phospho Y216 + Y279) with ab68476 at 1/1000 dilution. 5% NFDM/TBST was used as the diluting and blocking buffer. ab97051 Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated was used as the secondary antibody at 1/100000 dilution. Exposure time: 3 minutes.
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All lanes : Anti-GSK3 beta (phospho Y216) + GSK3 alpha (phospho Y279) antibody [EPR933Y] (ab68476) at 1/2000 dilution (unpurified)
Lane 1 : HEK-293 (Human epithelial cell line from embryonic kidney) cell lysate, cells untreated.
Lane 2 : HEK-293 cell lysate, cells treated with AP.
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : HRP conjugated Goat anti-rabbit at 1/2000 dilution
Predicted band size: 47-52 kDa
Observed band size: 47-52 kDa
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (40)
ab68476 has been referenced in 40 publications.
- Wen J et al. Pharmacological suppression of glycogen synthase kinase-3 reactivates HIV-1 from latency via activating Wnt/β-catenin/TCF1 axis in CD4+ T cells. Emerg Microbes Infect 11:391-405 (2022). PubMed: 34985411
- Shimizu Y et al. GSK3 inhibition circumvents and overcomes acquired lorlatinib resistance in ALK-rearranged non-small-cell lung cancer. NPJ Precis Oncol 6:16 (2022). PubMed: 35301419
- Brody AH et al. Alzheimer risk gene product Pyk2 suppresses tau phosphorylation and phenotypic effects of tauopathy. Mol Neurodegener 17:32 (2022). PubMed: 35501917
- Zhao C et al. GSK3β palmitoylation mediated by ZDHHC4 promotes tumorigenicity of glioblastoma stem cells in temozolomide-resistant glioblastoma through the EZH2-STAT3 axis. Oncogenesis 11:28 (2022). PubMed: 35606353
- Kasacka I et al. Wnt/β-catenin signaling in the adrenal glands of rats in various types of experimental hypertension. Endocr Connect 11:N/A (2022). PubMed: 35904223