Phospho-AKT1 (T450) Recombinant Rabbit Monoclonal Antibody [SD08-12]
cat.: ET1612-73
| Product Type: | Recombinant Rabbit monoclonal IgG, primary antibodies |
|---|---|
| Species reactivity: | Human, Mouse, Rat |
| Applications: | WB, IHC-P, IP, IF-Cell, FC |
| Clonality: | Monoclonal |
| Clone number: | SD08-12 |
| Form: | Liquid |
| Storage condition: | Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles. |
| Storage buffer: | 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide. |
| Concentration: | 1ug/ul |
| Purification: | Protein A affinity purified. |
| Molecular weight: | Predicted band size: 56 kDa |
| Isotype: | IgG |
| Immunogen: | Synthetic phospho-peptide corresponding to residues surrounding Thr450 of human AKT1. |
| Positive control: | A549 cell lysate, MCF7 cell lysate, HeLa cell lysate, Jurkat cell lysate, C2C12 cell lysate, NIH/3T3 cell lysate, C6 cell lysate, PC-12 cell lysate, human breast cancer tissue, mouse kidney tissue, rat kidney tissue, human colon carcinoma tissue, mouse lung tissue, MCF7, C2C12, C6. |
| Subcellular location: | Cytoplasm, Nucleus, Cell membrane. |
| Recommended Dilutions:
WB IHC-P IP IF-Cell FC |
1:500-1:2,000 1:50-1:200 Use at an assay dependent concentration. 1:100 1:1,000 |
| Uniprot #: | SwissProt: P31749 Human | P31750 Mouse | P47196 Rat |
| Alternative names: | AKT 1 AKT AKT1 AKT1_HUMAN MGC99656 PKB PKB-ALPHA PRKBA Protein Kinase B Alpha Protein kinase B Proto-oncogene c-Akt RAC Alpha RAC RAC-alpha serine/threonine-protein kinase RAC-PK-alpha |
Images
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Fig1:
Western blot analysis of Phospho-AKT1 (T450) on different lysates with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/1,000 dilution. Lane 1: A549 cell lysate Lane 2: MCF7 cell lysate Lane 3: HeLa cell lysate Lane 4: Jurkat cell lysate Lane 5: C2C12 cell lysate Lane 6: NIH/3T3 cell lysate Lane 7: C6 cell lysate Lane 8: PC-12 cell lysate Lane 9: MCF7 cell lysate, the membrane treated with λpp for 1 hour Lane 10: C2C12 cell lysate, the membrane treated with λpp for 1 hour Lane 11: C6 cell lysate, the membrane treated with λpp for 1 hour Lysates/proteins at 20 µg/Lane. Predicted band size: 54 kDa Observed band size: 54 kDa Exposure time: 3 minutes; ECL: K1802; 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 (ET1612-73) at 1/1,000 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. |
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Fig2:
Immunohistochemical analysis of paraffin-embedded human breast cancer tissue with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/200 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (ET1612-73) at 1/200 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. |
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Fig3:
Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/200 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (ET1612-73) at 1/200 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. |
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Fig4:
Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/200 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (ET1612-73) at 1/200 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. |
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Fig5:
Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/50 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 (ET1612-73) at 1/50 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. |
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Fig6:
Immunohistochemical analysis of paraffin-embedded mouse lung tissue with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/200 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 (ET1612-73) at 1/200 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. |
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Fig7:
Immunocytochemistry analysis of MCF7 cells labeling Phospho-AKT1 (T450) with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. |
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Fig8:
Flow cytometric analysis of MCF7 cells labeling Phospho-AKT1 (T450). Cells were fixed and permeabilized. Then stained with the primary antibody (ET1612-73, 1/1,000) (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). |
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Fig9:
Immunocytochemistry analysis of C2C12 cells labeling Phospho-AKT1 (T450) with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. |
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Fig10:
Flow cytometric analysis of C2C12 cells labeling Phospho-AKT1 (T450). Cells were fixed and permeabilized. Then stained with the primary antibody (ET1612-73, 1/1,000) (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). |
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Fig11:
Immunocytochemistry analysis of C6 cells labeling Phospho-AKT1 (T450) with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-AKT1 (T450) antibody (ET1612-73) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. |
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Fig12:
Flow cytometric analysis of C6 cells labeling Phospho-AKT1 (T450). Cells were fixed and permeabilized. Then stained with the primary antibody (ET1612-73, 1/1,000) (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). |
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