Overview

  • Product name
    NF kappaB p65 (pS536 + Total) ELISA Kit
  • Detection method
    Colorimetric
  • Precision
    Intra-assay
    Sample n Mean SD CV%
    (pS536) 6 3.4%
    (Total) 6 2.8%
    Inter-assay
    Sample n Mean SD CV%
    (pS536) 3 10.3%
    (Total) 3 8.7%
  • Sample type
    Cell Lysate, Tissue Homogenate
  • Assay type
    Semi-quantitative
  • Sensitivity
    500 pg/ml
  • Assay time
    1h 30m
  • Assay duration
    One step assay
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Rat
  • Product overview

    Abcam’s NF?B p65 (pS536) and NF?B p65 (Total) in vitro SimpleStep ELISA™ (Enzyme-Linked Immunosorbent Assay) kit is designed for the semi-quantitative measurement of NF?B p65 (pS536) and Total NF?B p65 protein in Human and mouse cells.

    The SimpleStep ELISA™ employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.

  • Notes

    Estimated sensitivity: Phospho-NF-κB p65 (Ser536): 5 µg/mL (tested with HeLa), Total NF-κB p65: 5 µg/mL (tested with HeLa)
    Range: Phospho-NF-κB p65 (Ser536):  6-600 µg/mL, Total NF-κB p65: 6-600 µg/mL

  • Tested applications
    Suitable for: Sandwich ELISAmore details
  • Platform
    Microplate

Properties

Applications

Our Abpromise guarantee covers the use of ab176663 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

  • Example of a typical NFĸB p65 (pS536) and NFĸB p65 (Total) cell lysate dilution series. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.

  • Linearity of dilution in representative sample matrices. Cellular lysates were prepared at 3 concentrations in common media containing 1X Cell Extraction Buffer PTR. Data from duplicate measurements of NFĸB p65 (pS536) are normalized and plotted.

  • Cell line analysis for Total NFĸB p65 from 200 µg/mL preparations of cell extracts. Data from triplicate measurements (mean +/- SD) are plotted and compared to 1X Cell Extraction Buffer PTR (zero).

  • Induction of NFĸB p65 (pS536) phosphorylation in MCF-7 cells in response to TNFα treatment. HeLa cells were cultured in 96-well tissue culture plates, and treated (10 min) with a dose-range of TNFα before cell lysis. Data from quadruplicate measurements of NFĸB p65 (pS536) are plotted and compared against total NFĸB p65 protein levels. Comparative NFĸB p65 (pS536) and NFĸB p65 (Total) data also shown by Western Blot.

Protocols

References

This product has been referenced in:
  • Song L  et al. c-Kit modifies the inflammatory status of smooth muscle cells. PeerJ 5:e3418 (2017). ELISA ; Mouse . Read more (PubMed: 28626608) »
  • Kroetz DN  et al. Type I Interferon Induced Epigenetic Regulation of Macrophages Suppresses Innate and Adaptive Immunity in Acute Respiratory Viral Infection. PLoS Pathog 11:e1005338 (2015). Sandwich ELISA ; Human . Read more (PubMed: 26709698) »
See all 2 Publications for this product

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

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