AKT antibody

• Catalog Number: orb344419
• Category: Antibodies
• Description: AKT antibody
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 14E5.16C8.25F6
• Tested applications: ELISA, FC, IHC, WB
• Reactivity: Human, Mouse, Primate, Rat
• Isotype: IgG
• Immunogen: Anti-AKT Antibody was produced by repeated immunizations in mice with a synthetic peptides corresponding to residues internal to the human AKT1, AKT2, and AKT3 proteins.
• Concentration: 1.0 mg/mL
• Dilution range: ELISA: 1:20,000, FC: User Optimized, IHC: 20 µg/mL, WB: 1:500 - 1:3,000
• Form/Appearance: Liquid (sterile filtered)
• Purity: Anti-AKT Antibody was purified by Protein A chromatography. This antibody is specific for human and mouse AKT protein. A BLAST analysis was used to suggest cross-reactivity with AKT1, AKT2, and AKT3 from human, mouse, rat and chimpanzee sources based on 100% homology with the immunizing sequence. Cross-reactivity with AKT1, 2, 3 was determined with Western Blot. Cross reactivity of AKT from other sources has not been determined.
• Conjugation: Unconjugated
• UniProt ID: P31749
• NCBI: 62241011
• Storage: Store vial at -20° C prior to opening. Aliquot contents and freeze at -20° C or below for extended storage. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use.
• Buffer/Preservatives: 0.01% (w/v) Sodium Azide
• Alternative names: mouse anti-AKT Antibody, RAC-PK-alpha, Protein kin
• Note: For research use only
• Application notes: Mouse Anti-AKT Antibody is tested for ELISA, SDS-PAGE, and western blotting. This antibody is suitable for immunohistochemistry and immunoprecipitation. Expect a band approximately 54 - 56 kDa in size corresponding to AKT protein by western blotting in the appropriate cell lysate or extract. Specific conditions for reactivity should be optimized by the end user. For immunohistochemistry we recommend the use of fresh frozen tissues. Attempts at staining paraffin-embedded formalin fixed tissues were negative. No pre-treatment of sample is required.
• Expiration Date: 12 months from date of receipt.

ELISA of Mouse Anti-AKT Antibody. Antigen: HCT-116 cell line (p/n orb348712). Coating amount: Confluent in the 96 well plate. Primary antibody: AKT (top) or GAPDH (bottom) antibody at 2 µg/mL. Dilution series: Primary and Secondary Antibodies 2-fold. Mid-point concentration: N/A. Secondary antibody: DyLight™ 680 donkey secondary antibody and DyLight™ 800 goat secondary antibody starting at 1:1000. Substrate: None.

SDS PAGE of Mouse anti-AKT Monoclonal Antibody. Lane 1: Reduced Mouse anti-AKT Monoclonal Antibody. Lane M: 3 µL Opal Prestained Marker. Lane 2: Non-Reduced Mouse anti-AKT Monoclonal Antibody. Load: 1 µg per lane. Predicted/Observed size: Non-Reduced at 160 kDa/Observed at 245 kDa; Reduced at 55, 25 kDa. Non-reduced migrates slightly higher.

Western Blot of Mouse anti-AKT antibody. Lane 1: NIH/3T3 cell lysates (p/n orb348714). Lane 2: PDGF stimulated NIH/3T3 cell lysates (p/n orb348723). Load: 10 µg per lane. Primary antibody: Anti-AKT antibody at 1:400 for overnight at 4°C. Secondary antibody: Goat-anti-Mouse IgG HRP conjugated (p/n orb347385) was used at a 1:40000 dilution for 1 hr at 2-8°C with FemtoMax™ enhanced chemiluminescent reagent. Block: 5% BLOTTO (p/n orb348624) in TBS for 2hr at RT. Observed size: ~56 kDa for AKT.

Western Blot of Mouse anti-AKT Monoclonal Antibody. Lane 1: His-AKT1 Recombinant Protein. Lane 2: His-AKT2 Recombinant Protein. Lane 3: His-AKT3 Recombinant Protein. Load: 50 ng per lane. Primary antibody: Mouse anti-AKT Monoclonal Antibody at 1:1000 overnight at 4°C. Secondary antibody: HRP mouse secondary antibody at 1:40000 for 30 min at RT. Block: orb348637 for 30 min at RT. Predicted/Observed size: 54-56 kDa, 54-56 kDa for AKT1, AKT2, AKT3.