Overview

  • Product name

    Akt (pS473) + total Akt ELISA Kit
  • Detection method

    Colorimetric
  • Sample type

    Cell Lysate
  • Assay type

    Semi-quantitative
  • Assay time

    5h 00m
  • Assay duration

    Multiple steps standard assay
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Product overview

    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.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Microplate

Properties

Associated products

Applications

Our Abpromise guarantee covers the use of ab126433 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Sandwich ELISA Use at an assay dependent concentration.

Images

  • 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.

Protocols

References

This product has been referenced in:

  • 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). Read more (PubMed: 30905432) »
  • Riddle MR  et al. Insulin resistance in cavefish as an adaptation to a nutrient-limited environment. Nature 555:647-651 (2018). Read more (PubMed: 29562229) »
See all 5 Publications for this product

Customer reviews and Q&As

Filter by Ratings

Compatibility with Porcine cell (LLC-PK1)

Average Good 4/5 (Ease of Use)
Abreviews
I have used this product to measure the increase in the phosphorylation of AKT when treated with TGF-beta. I found the values to be consistent across the control and TGF-beta treated groups. The phosphorylation of AKT also increased on treatment with TGF-beta as the literature suggests. I feel this product to be reliable in the porcine model as well.
However, the problem I had with the kit was with the standard curve. Firstly, the lyophillized cell lysate that was provided precipitated and I could not obtain a proper standard curve. Moreover, I feel that it would be a good idea for abcam to quantitate the amount of AKT present in the cell lysate they provide so that the results may be expressed as (amount of pAKT/ amount of total AKT) instead of the (absorbance of pAKT/ absorbance of total AKT) as it is being done now.

Abcam user community

Verified customer

Submitted Sep 18 2013

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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