Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Flow cytometry [1]
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Validation data
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- Product number
- 701052 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Phospho-AKT1 (Thr308) Recombinant Rabbit Monoclonal Antibody (B18H12L21)
- Antibody type
- Monoclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- B18H12L21
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of AKT (pT308) was performed by loading 30 µg of U-87 MG cell lysate (lane 1) using Novex®NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5% skim milk for 1 hour at room temperature. AKT (pT308) was detected at ~70 kDa using AKT (pT308) Recombinant Rabbit Monoclonal Antibody (Product # 701052) at a 1:1000 dilution in 2.5% skim milk at 4°C overnight on a rocking platform. To confirm specificity, competition was performed with the phosphopeptide (10 µg/mL) (lane 2). Detection was performed using an HRP-conjugated Goat anti-Rabbit secondary antibody (Product # G-21234) at a 1:5000 dilution and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of Phospho-AKT pThr308 in whole cell extracts of U87-MG cells using a Phospho-AKT pThr308 recombinant rabbit monoclonal antibody (Product # 701052) at a dilution of 1 µg/mL. To confirm specificity, competition was performed by preincubation with the phosphopeptide to inhibit antibody binding (lane 2). Samples were detected using chemiluminescence (ECL). Results show a band at ~70kDa.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of Phospho-AKT pThr308 in serum-starved HeLa cells treated with insulin (100 ng/mL, 15 min) using a Phospho-AKT pThr308 recombinant rabbit monoclonal antibody (Product # 701052) followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody (green) (Image A). Nuclei were stained using DAPI (Image B) and actin stained with Alexa Fluor 594 phalloidin (red) (image C). Image D is a composite image showing nuclear localization of phosphorylated AKT and Image E is a composite image of cells showing competition with the phospho AKT (pT308) peptide.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow cytometry analysis of AKT (pT308) was performed on Jurkat cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0. 25% Tritonª X-100 for 20 minutes, and blocked with 5% BSA for 1 hour at room temperature. Cells were labeled with ABfinityª AKT (pT308) recombinant rabbit monoclonal antibody (Product # 701052, red histogram) or with rabbit isotype control (pink histogram) at a dilution of 1:250 in 2.5% BSA. After incubation at room temperature for 3 hours, the cells were labeled with Alexa Fluor¨ 488 goat anti-Rabbit Secondary antibody (Product # A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune¨ Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.