Key features and details
- Sample type: Cell Lysate
- Detection method: Colorimetric
- Assay type: Semi-quantitative
- Reacts with: Mouse, Rat, Human
Product nameAkt (pS473) + total Akt ELISA Kit
Sample typeCell Lysate
Assay time5h 00m
Assay durationMultiple steps standard assay
Species reactivityReacts with: Mouse, Rat, Human
ab126433 is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in cell lysates. By determining phosphorylated Akt protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blot analysis.
This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of phospho-Akt (Ser473) and total Akt in human, mouse and rat cell lysates (help normalize the results of phospho-Akt from different cell lysate being compared). A pan Akt antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and Akt present in a sample is bound to the wells by the immobilized antibody. The wells are washed and anti-phospho-Akt (Ser473) or anti-pan-Akt is used to detect phosphorylated or total Akt. After washing away unbound antibody, HRP-conjugated anti-rabbit IgG is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of Akt (Ser473) or total Akt bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.
Get higher sensitivity in only 90 minutes with AKT 1/2/3 pS473 + AKT1 ELISA Kit (ab176657) from our SimpleStep ELISA® range.
Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.
Storage instructionsStore at -20°C. Please refer to protocols.
Components 1 x 96 tests 500X HRP-conjugated anti-rabbit IgG 1 x 25µl 20X Wash Buffer Concentrate 1 x 25ml 2X Cell Lysis Buffer 1 x 5ml 5X Assay Diluent 1 x 15ml Akt Microplate (12 strips x 8 wells) coated with anti-pan Akt antibody 1 unit Detection Antibody Akt (Ser473): rabbit anti-phospho-Akt (Ser473) 1 vial Detection Antibody Akt: rabbit anti-pan-Akt 1 vial Positive Control: lyophilized NIH3T3 cell lysate 1 vial Stop Solution 1 x 8ml TMB One-Step Substrate Reagent 1 x 12ml
- Epigenetics and Nuclear Signaling
- Domain Families
- HLH / Leucine Zipper
- HLH / Leucine Zipper
Cellular localizationAKT1/2/3: Cytoplasm. Membrane. Membrane-associated after cell stimulation leading to its translocation. AKT1 (total): Cytoplasm. Nucleus. Cell membrane. Nucleus after activation by integrin-linked protein kinase 1 (ILK1). Nuclear translocation is enhanced by interaction with TCL1A. Phosphorylation on Tyr-176 by TNK2 results in its localization to the cell membrane where it is targeted for further phosphorylations on Thr-308 and Ser-473 leading to its activation and the activated form translocates to the nucleus.
- AKT 1
MCF7 cells were stimulated for five minutes with 1 ug x mL-1 of insulin (ab123768). Cell lysates were assessed for total Akt and Akt(pS473), shown as OD (450 nm) after background signal was subtracted (duplicates +/- SD).
The NIH3T3 cells were treated with recombinant human PDGF for 10 minutes to induce phosphorylation of Akt. Serial dilutions of lysates were analyzed in this ELISA.
NIH3T3 cells were treated or untreated with recombinant human PDGF for 10 min. Cell lysates were analyzed by Western Blot.
NIH3T3 cells were treated or untreated with recombinant human PDGF for 10 min. Cell lysates were analyzed using this phosphoELISA.
A431 cells were treated with recombinant human EGF at 37°C for 20 min. Solubilize cells at 4 x 107 cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA.
ab126433 has been referenced in 5 publications.
- Guan Y et al. Chloride channel-3 is required for efficient tumour cell migration and invasion in human cervical squamous cell carcinoma. Gynecol Oncol 153:661-669 (2019). PubMed: 30905432
- Riddle MR et al. Insulin resistance in cavefish as an adaptation to a nutrient-limited environment. Nature 555:647-651 (2018). PubMed: 29562229
- Ding WZ et al. MicroRNA-497 regulates cell proliferation in hepatocellular carcinoma. Oncol Lett 11:1081-1088 (2016). WB . PubMed: 26893696
- Gerdes JM et al. Ciliary dysfunction impairs beta-cell insulin secretion and promotes development of type 2 diabetes in rodents. Nat Commun 5:5308 (2014). Sandwich ELISA ; Mouse . PubMed: 25374274
- Arroba AI et al. Loss of protein tyrosine phosphatase 1B increases IGF-I receptor tyrosine phosphorylation but does not rescue retinal defects in IRS2-deficient mice. Invest Ophthalmol Vis Sci 54:4215-25 (2013). PubMed: 23702782