Anti-Phospho-GSK3B (Ser9) Polyclonal Antibody is a Rabbit antibody targeting Phospho-GSK3B (Ser9). Anti-Phospho-GSK3B (Ser9) Polyclonal Antibody can be used in WB,IHC-P,IHC-Fr,ICC/IF,IF,FCM.

IgG

Human,Mouse,Rat

1. Sample: Testis (Rat) Lysate at 30 μg
Primary: Anti-phospho-GSK-3 Beta (Ser9) (TMAB-01416) at 1:200 dilution;
Secondary: HRP conjugated Goat Anti-Rabbit Igg (secondary antibody) at 1: 5000 dilution;
Predicted band size: 47 kDa
Observed band size: 49 kDa
2. The blue histogram is unstained cells (A549 cells).
The Orange histogram is cells stained with Rabbit IgG/FITC.
The green histogram is cells stained with Rabbit Anti-phospho-GSK-3 Beta (Ser9)/FITC Conjugated antibody (TMAB-01416-FITC).
Isotype control: Cell lines treated with Rabbit IgG/FITC instead of the primary antibody to confirm that primary antibody binding is specific. Concentration: 5 μL in 100 μL 1 X PBS containing 0.5% BSA.
3. Tissue/cell: rat kidney tissue; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer (0.01M, pH6.0), Boiling bathing for 15 min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30 min; Blocking buffer (normal goat serum) at 37°C for 20 min;
Incubation: Anti-phospho-GSK-3 Beta (Ser9) Polyclonal Antibody, Unconjugated (TMAB-01416) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody and DAb staining.
4. Paraformaldehyde-fixed, paraffin embedded (Mouse placenta); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 min; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (Beta (Ser9)) Polyclonal Antibody, Unconjugated (TMAB-01416 p-GSK-3) at 1:500 overnight at 4°C, followed by a conjugated secondary for 20 min and DAB staining.
5. Blank control: A431. Primary Antibody (green line): Rabbit Anti-phospho-GSK-3 Beta (Ser9) antibody (TMAB-01416)
Dilution: 1 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-AF488
Dilution: 1 μg/test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 90% ice-cold methanol for 20 min at-20°C. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature.
6. Tissue/cell: Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (phospho-GSK-3 Beta (Ser9)) polyclonal Antibody, Unconjugated (TMAB-01416) 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nucleus.
7. Sample:
Lane 1: Mouse Cerebrum tissue lysates
Lane 2: Rat Cerebrum tissue lysates
Primary: Anti-phospho-GSK-3 Beta (Ser9) (TMAB-01416) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 47 kDa
Observed band size: 47 kDa

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; ICC/IF: 1:100; IF: 1:100-500; FCM: 1ug/Test

Polyclonal

KLH conjugated Synthesised phosphopeptide: human GSK-3 Beta around the phosphorylation site of Ser9

Human

GSK3B

Glycogen synthase kinase-3 beta

Integration of energy metabolism,Metabolism of carbohydrates,Diabetes associated,Hypertrophy,Carbohydrate metabolism,Integration of energy,Cancer,Diabetes,Heart disease,Notch Pathway,Other Kinases,Cytoplasmic,Cytoplasmic

Constitutively active protein kinase that acts as a negative regulator in the hormonal control of glucose homeostasis, Wnt signaling and regulation of transcription factors and microtubules, by phosphorylating and inactivating glycogen synthase (GYS1 or GYS2), EIF2B, CTNNB1/beta-catenin, APC, AXIN1, JUN, NFATC1/NFATC, MAPT/TAU and MACF1. Requires primed phosphorylation of the majority of its substrates. In skeletal muscle, contributes to insulin regulation of glycogen synthesis by phosphorylating and inhibiting GYS1 activity and hence glycogen synthesis. May also mediate the development of insulin resistance by regulating activation of transcription factors. Regulates protein synthesis by controlling the activity of initiation factor 2B (EIF2BE/EIF2B5) in the same manner as glycogen synthase. In Wnt signaling, GSK3B forms a multimeric complex with APC, AXIN1 and CTNNB1/beta-catenin and phosphorylates the N-terminus of CTNNB1 leading to its degradation mediated by ubiquitin/proteasomes. Phosphorylates JUN at sites proximal to its DNA-binding domain, thereby reducing its affinity for DNA. Phosphorylates NFATC1/NFATC on conserved serine residues promoting NFATC1/NFATC nuclear export, shutting off NFATC1/NFATC gene regulation, and thereby opposing the action of calcineurin. Phosphorylates MAPT/TAU on 'Thr-548', decreasing significantly MAPT/TAU ability to bind and stabilize microtubules. MAPT/TAU is the principal component of neurofibrillary tangles in Alzheimer disease. Plays an important role in ERBB2-dependent stabilization of microtubules at the cell cortex. Phosphorylates MACF1, inhibiting its binding to microtubules which is critical for its role in bulge stem cell migration and skin wound repair. Probably regulates NF-kappa-B (NFKB1) at the transcriptional level and is required for the NF-kappa-B-mediated anti-apoptotic response to TNF-alpha (TNF/TNFA). Negatively regulates replication in pancreatic beta-cells, resulting in apoptosis, loss of beta-cells and diabetes. Phosphorylates MUC1 in breast cancer cells, decreasing the interaction of MUC1 with CTNNB1/beta-catenin. Is necessary for the establishment of neuronal polarity and axon outgrowth. Phosphorylates MARK2, leading to inhibit its activity. Phosphorylates SIK1 at 'Thr-182', leading to sustain its activity.