RPS6 pSer240/244 + Total PhosphoTracer ELISA Kit (ab119668)

Overview

  • Product nameRPS6 pSer240/244 + Total PhosphoTracer ELISA Kit
  • Detection methodFluorescent
  • Tests
    1 x 96 well plate
  • Sample type
    Cell culture extracts
  • Assay typeSemi-quantitative
  • Assay time
    2h 0m
  • Assay durationOne step assay
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Rat
  • Product overview

    PhosphoTracer assays use a traditional immuno-sandwich format, but with a major difference both the analyte and the assay reagents are added to the PhosphoTracer assay microplate at the same time. After a short incubation period, unbound assay reagents and analytes are washed away, and immuno-complexes containing both antibodies are detected. The process can take as little as 60 minutes to complete.

    PhosphoTracer kits also allow a higher degree of assay flexibility. In contrast to other ELISA formats, no antibodies are present on the assay microplate itself, so assays for several different targets can be performed in different wells on the same microplate. Simply mix the lysate with your target reagents of choice, using the microplate configuration of your choice.

    A whole new way of performing cellular assays, PhosphoTracer takes the hard work out of running a standard ELISA, while still giving the high quality results expected from a sandwich immunoassay. Fully self-contained kits are supplied in convenient 96-well packs. Simple to use and highly sensitive PhosphoTracer kits are designed to get results, fast.

     

    Abcam’s PhosphoTracer RPS6 (pSer240/244) assay kits detect endogenous levels of RPS6 (GenBank Accession NP_001001) in cellular lysates. The phospho-RPS6 assay detects RPS6 only when phosphorylated at Ser240/244. The total RPS6 assay detects RPS6 irrespective of phosphorylation status.



    The substrate used with the HRP conjugated detection antibody is a combination of 10-Acetyl-3,7-dihydroxyphenoxazine (ADHP) (wavelength exc/em = 530-540nm / 590-600nm), a highly sensitive and stable substrate for HRP) and ADHP Dilution Buffer (a stabilized H2O2 solution). Learn more about the fluorogenic substrate, ADHP.

  • Notes

    Sensitivity: Phospho-RPS6: 0.1 ug protein/ml (tested in MCF7 cell lysate), RPS6: 1 ug protein/ml (tested in MCF7 cell lysate).

    Range: Phospho-RPS6: 0.1-250 ug protein/ml (tested in MCF7 cell lysate), RPS6: 1-250 ug protein/ml (tested in MCF7 cell lysate).

  • Tested applicationsSandwich ELISAmore details
  • PlatformMicroplate

Properties

Applications

Our Abpromise guarantee covers the use of ab119668 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. Phospho-RPS6: Tested in MCF7, PC3, NIH3T3; RPS6: Tested in MCF7, PC3, NIH3T3, A549, HEK293, HeLa, U2OS

RPS6 pSer240/244 + Total PhosphoTracer ELISA Kit images

  • Using Western blot, RPS6 phosphorylation at Ser240/244 is detected in insulin-treated MCF7 cells (+), compared with MCF7 cells treated with a combination of rapamycin, UCN-01 and U0126 (-), whereas no change in total RPS6 levels was observed.
  • Using the RPS6 assay kits, RPS6 phosphorylation at Ser240/244 is detected in insulin-treated MCF7 cells (+), compared with MCF7 cells treated with a combination of rapamycin, UCN-01 and U0126 (-), whereas no change in total RPS6 levels was observed.
  • MCF7 cells were seeded at 40K cells/well in medium containing 10% FBS in a 96 well microplate, and cultured overnight. The following day the culture medium was removed, and the cells were treated with various concentrations of rapamycin diluted in serum-free medium for 60 mins. The medium was removed from the wells, and the cells were lysed with 120 µL/well Lysis Mix, with shaking for 10 min. 50 µL of lysate was transferred to replicate wells of a PhosphoTracer assay plate, and Antibody Mix specific for either phospho RPS6 or total RPS6 (50 µL/well) was added to the lysates. The plates were incubated for 1 hr at room temp, with shaking. Plates were washed and Substrate Mix was added. The plates were covered in foil and incubated for 10 min with shaking. Signal in the wells was determined using a plate reader.

Protocols

References for RPS6 pSer240/244 + Total PhosphoTracer ELISA Kit (ab119668)

ab119668 has not yet been referenced specifically in any publications.

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