Phospho-mTOR (Ser2481) Rabbit Polyclonal Antibody

• Catalog Number: orb6453
• Category: Antibodies
• Description: Phospho-mTOR (Ser2481) Rabbit Polyclonal Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, ICC, IF, IHC-Fr, IHC-P, WB
• Predicted Reactivity: Canine, Equine, Porcine, Rabbit, Sheep
• Reactivity: Human, Mouse, Rat
• Isotype: IgG
• Immunogen: KLH conjugated Synthesised phosphopeptide derived from human mTOR around the phosphorylation site of Ser2481 IH(p-S)FI
• Concentration: 1mg/ml
• Dilution range: WB=1:500-2000, IHC-P=1:100-500, IHC-F=1:100-500, ICC/IF=1:100, IF=1:100-500, Flow-Cyt=1μg /test
• Form/Appearance: Liquid
• Conjugation: Unconjugated
• MW: 289 kDa
• Target: MTOR
• UniProt ID: P42345
• RRID: AB_10961108
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Buffer/Preservatives: 0.01M TBS (pH7.4) with 1% rAlbumin, 0.02% Proclin300 and 50% Glycerol.
• Alternative names: mTOR (phospho S2481); p-mTOR (phospho S2481); dJ57
• Note: For research use only
• Expiration Date: 12 months from date of receipt.

A431 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-mTOR (Ser2481)) polyclonal Antibody, Unconjugated (orb6453) 1:100, 90 minutes at 37°C, followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nuclei.

Blank control (blue line): Hela (fixed with 70% ethanol (Overnight at 4°C) and then permeabilized with 90% ice-cold methanol for 30 min on ice), Primary Antibody (green line): Rabbit Anti-Phospho-mTOR (Ser2481) antibody (orb6453), dilution: 1 µg/10^6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody (white blue line): Goat anti-rabbit IgG-FITC, dilution: 1 µg/Test.

Blank control: A431. Primary Antibody (green line): Rabbit Anti-Phospho-mTOR (Ser2481) antibody (orb6453), dilution: 1 ug/Test, Secondary Antibody: Goat anti-rabbit IgG-AF488, dilution: 0.5 ug/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. Acquisition of 20000 events was performed.

Blank control: mouse splenocytes (blue), Isotype Control Antibody: Rabbit IgG (orange), Secondary Antibody: Goat anti-rabbit IgG-FITC (white blue), dilution: 1:100 in 1X PBS containing 0.5% BSA, Primary Antibody, dilution: 5 µl in 100 µl 1X PBS containing 0.5% BSA (green).

Paraformaldehyde-fixed, paraffin embedded (mouse brain tissue), Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min, Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes, Blocking buffer (normal goat serum) at 37°C for 30 min, Antibody incubation with (Phospho-mTOR (Ser2481)) Polyclonal Antibody, Unconjugated (orb6453) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat brain), Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min, Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes, Blocking buffer (normal goat serum) at 37°C for 30 min, Antibody incubation with (Phospho-mTOR (Ser2481)) Polyclonal Antibody, Unconjugated (orb6453) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.

Sample: Hepg2 (Human) Cell Lysate at 30 ug, Primary: Anti-Phospho-mTOR (Ser2481) (orb6453) at 1/300 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 289 kD, Observed band size: 289 kD.

Sample: Jurkat (Human) Cell Lysate at 30 ug, MCF-7 (Human) Cell Lysate at 30 ug, Primary: Anti-Phospho-mTOR (Ser2481) (orb6453) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 289 kD, Observed band size: 289 kD.

Tissue/Cell: rat brain 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-mTOR (Ser2481) Polyclonal Antibody, Unconjugated (orb6453) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody and DAB staining.