STAT5 Total PhosphoTracer ELISA Kit (ab119653)

Overview

  • Product nameSTAT5 Total PhosphoTracer ELISA Kit
  • Detection methodFluorescent
  • Tests
    1 x 96 well plate
  • Sample type
    Cell culture extracts
  • Assay typeSemi-quantitative
  • Sensitivity
    10 µg protein/ml
  • Range
    10 µg protein/ml - 1000 µg protein/ml
  • Assay time
    2h 0m
  • Assay durationOne step assay
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Mouse, 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 phospho-STAT5 assay kits detect endogenous levels of STAT5A and STAT5B (GenBank Accessions NP_003143 and NP_036580) in cellular lysates. Total STAT5 assay kits detect STAT5 irrespective of phosphorylation.



    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.

  • Tested applicationsSandwich ELISAmore details
  • PlatformMicroplate

Properties

Applications

Our Abpromise guarantee covers the use of ab119653 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. Tested in A431, THP1, NIH3T3

STAT5 Total PhosphoTracer ELISA Kit images

  • Using Western blot, a significant stimulation of STAT5 phosphorylation at Tyr694/699 is detected in A431 cells treated with EGF for 10 minutes (+) compared with cells treated with AG1478 (-).
  • Using the phospho-STAT5 assay kit, a significant stimulation of STAT5 phosphorylation at Tyr694/699 is detected in A431 cells treated with EGF for 10 minutes (+) compared with cells treated with AG1478 (-).
  • Day 1: A431 cells were seeded at 25K/well in a 96 well tissue culture plate in medium containing 10% FBS. Day 2: The cells were stimulated with various concentrations of EGF for 15 minutes. The media was removed, and the cells were lysed with 125 µL/well freshly prepared Lysis Mix, with shaking for 10 min. Lysates (50 µL) were transferred to a PhosphoTracer assay plate and analyzed for either phospho STAT5 using the standard PhosphoTracer protocol. Briefly, Antibody Mix specific for phospho-STAT5 (50 µL/well), was added to the lysate, and the plate was incubated for 1 hr at room temp, with shaking. The plate was washed and Substrate Mix was added to the wells. The plate was covered in foil and incubated for 10 min with shaking. Signal in the wells was determined using a plate reader.

Protocols

References for STAT5 Total PhosphoTracer ELISA Kit (ab119653)

ab119653 has not yet been referenced specifically in any publications.

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While we have not validated our individual primary antibodies as sELISA pairs, we do have three PhosphoTracer ELISA kits that may interest you:
ab119652: PhosphoTracer STAT5 (pY694/699) ELISA Kit
ab119653: Ph...

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"