Phospho-GSK3 (alpha + beta) (Y216 + Y279) Recombinant Rabbit Monoclonal Antibody [SY26-05] - BSA and Azide free
Catalog# HA750127
Phospho-GSK3 (alpha + beta) (Y216 + Y279) Recombinant Rabbit Monoclonal Antibody [SY26-05] - BSA and Azide free
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WB
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IF-Cell
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IHC-P
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IP
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IF-Tissue
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FC
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Human
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Mouse
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Rat
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unconjugated
Overview
Product Name
Phospho-GSK3 (alpha + beta) (Y216 + Y279) Recombinant Rabbit Monoclonal Antibody [SY26-05] - BSA and Azide free
Antibody Type
Recombinant Rabbit monoclonal Antibody
Immunogen
Synthetic phospho-peptide corresponding to residues surrounding Tyr216 and 279 of human GSK3(alpha+beta).
Species Reactivity
Human, Mouse, Rat
Validated Applications
WB, IF-Cell, IHC-P, IP, IF-Tissue, FC
Target Molecular Weight
Predicted band size: 51 kDa
Positive Control
293T cell lysate, SH-SY5Y cell lysate, HeLa cell lysate, HepG2 cell lysate, MCF7 cell lysate, NIH/3T3 cell lysate, PC-12 cell lysate, mouse brain tissue lysate, rat brain tissue lysate, mouse cerebellum tissue lysates, A431, Hela, MCF-7, rat brain tissue, rat hippocampus tissue, mouse brain tissue, human thyroid carcinoma tissue.
Conjugation
unconjugated
Clone Number
SY26-05
Product Features
Form
Liquid
Concentration
1mg/ml
Storage Instructions
Store at 2-8℃. Avoid freeze.
Storage Buffer
1*PBS (pH7.4).
Isotype
IgG
Purification Method
Protein A affinity purified.
Application Dilution
Target
Function
Glycogen synthase kinase-3α (GSK-3α) and GSK-3β are highly similar isoforms of serine/ threonine kinases that regulate metabolic enzymes and transcription factors, which are responsible for coordinating processes such as glycogen synthesis and cell adhesion. GSK-3β activity is also required for nuclear activity of Rel dimers, which mediate an anti-apoptotic response to TNFα in mice. GSK-3 catalytic kinase activity is controlled through differential phosphorylation of serine/threonine residues, which have an inhibitory effect, and tyrosine residues, which have an activating effect. Growth factor stimulation of mammalian cells expressing GSK-3α and GSK-3β induces phosphorylation of Ser 21 and Ser 9, respectively, through a phosphatidylinositol 3-kinase (PI 3-K)-protein kinase B (PKB)-dependent pathway, thereby enhancing proliferative signals. Additionally, GSK-3 physically associates with cAMP-dependent protein kinase A (PKA), which phosphorylates Ser 21 of GSK-3α or Ser 9 of GSK-3β and inactivates both forms. GSK-3α/β is positively regulated by phosphorylation on Tyr 279 and Tyr 216, respectively. Activated GSK-3α/β participates in energy metabolism, neuronal cell development, and body pattern formation. Tyrosine dephosphorylation of GSK-3 is involved in its extracellular signal-dependent inactivation.
Background References
1. Xu R et al. The protease Omi regulates mitochondrial biogenesis through the GSK3 /PGC-1a pathway. Cell Death Dis 5:e1373 (2014).
2. Weigand S et al. Global quantitative phosphoproteome analysis of human tumor xenografts treated with a CD44 antagonist. Cancer Res 72:4329-39 (2012).
Sequence Similarity
Belongs to the protein kinase superfamily. CMGC Ser/Thr protein kinase family. GSK-3 subfamily.
Post-translational Modification
Phosphorylated by AKT1 at Ser-21: upon insulin-mediated signaling, the activated PKB/AKT1 protein kinase phosphorylates and desactivates GSK3A, resulting in the dephosphorylation and activation of GYS1. Activated by phosphorylation at Tyr-279.
Subcellular Location
Cytoplasm, Nucleus, Cell membrane.
UNIPROT
Synonyms
Factor A antibody
Glycogen synthase kinase 3 alpha antibody
Glycogen synthase kinase 3 beta antibody
GSK3 alpha antibody
GSK3 beta antibody
GSK3B antibody
Images
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☑ Cell treatment (CT)
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Western blot analysis of Phospho-GSK3 (alpha + beta) (Y216 + Y279) on different lysates with Rabbit anti-Phospho-GSK3 (alpha + beta) (Y216 + Y279) antibody (ET1607-54) at 1/5,000 dilution.
Lane 1: 293T cell lysate
Lane 2: SH-SY5Y cell lysate
Lane 3: HeLa cell lysate
Lane 4: HepG2 cell lysate
Lane 5: MCF7 cell lysate
Lane 6: NIH/3T3 cell lysate
Lane 7: PC-12 cell lysate
Lane 8: Mouse brain tissue lysate
Lane 9: Rat brain tissue lysate
Lane 10: 293T treated with λpp for 1 hour cell lysate
Lane 11: SH-SY5Y treated with λpp for 1 hour cell lysate
Lysates/proteins at 15 µg/Lane.
Predicted band size: 47/51 kDa
Observed band size: 47/51 kDa
Exposure time: 5 minutes;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1607-54) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:50,000 dilution was used for 1 hour at room temperature. -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
ICC staining of Phospho-GSK3 (alpha + beta) (Y216 + Y279) in A431 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1607-54, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue). -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
ICC staining of Phospho-GSK3 (alpha + beta) (Y216 + Y279) in Hela cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1607-54, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue). -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
ICC staining of Phospho-GSK3 (alpha + beta) (Y216 + Y279) in MCF-7 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1607-54, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue). -
☑ Cell treatment (CT)
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Western blot analysis of Phospho-GSK3(alpha+beta)(Y216+Y279) on mouse cerebellum tissue lysates.
Lane 1: mouse cerebellum tissue, whole cell lysate, 20ug/lane
Lane 2: mouse cerebellum tissue treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 20ug/lane
All lanes :
Anti-Phospho-GSK3(alpha+beta)(Y216+Y279) antibody (ET1607-54) at 1:500 dilution. Anti-GAPDH antibody (ET1601-4) at 1:10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (HA1001) at 1/200,000 dilution.
Predicted band size: 47/51 kDa
Observed band size: 47/51 kDa
Blocking and diluting buffer: 5% BSA.
Exposure time: 2 minutes -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-Phospho-GSK3 (alpha + beta) (Y216 + Y279) antibody (ET1607-54) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1607-54) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat hippocampus tissue with Rabbit anti-Phospho-GSK3 (alpha + beta) (Y216 + Y279) antibody (ET1607-54) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1607-54) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse brain tissue with Rabbit anti-Phospho-GSK3 (alpha + beta) (Y216 + Y279) antibody (ET1607-54) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1607-54) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded human thyroid carcinoma tissue with Rabbit anti-Phospho-GSK3 (alpha + beta) (Y216 + Y279) antibody (ET1607-54) at 1/800 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1607-54) at 1/800 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of HeLa cells labeling Phospho-GSK3 (alpha + beta) (Y216 + Y279).
Cells were fixed and permeabilized. Then stained with the primary antibody (ET1607-54, 1μg/mL) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black). -
This data was developed using ET1607-54, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of NIH/3T3 cells labeling Phospho-GSK3 (alpha + beta) (Y216 + Y279).
Cells were fixed and permeabilized. Then stained with the primary antibody (ET1607-54, 1μg/mL) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
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