SimpleStep

Human Cleaved PARP1 ELISA Kit (ab174441)

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Overview

  • Product name

    Human Cleaved PARP1 ELISA Kit

  • Detection method

    Colorimetric

  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Overall 5 6.6%
    Inter-assay
    Sample n Mean SD CV%
    Overall 3 8.5%

  • Sample type

    Cell culture extracts

  • Assay type

    Sandwich (quantitative)

  • Sensitivity

    5.8 µg/ml

  • Range

    7.8 µg/ml - 500 µg/ml

  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture media 59 43% - 73%
    Fetal Bovine Serum 65 52% - 85%
    Bovine Serum Albumin 75 57% - 94%

  • Assay time

    1h 30m

  • Assay duration

    One step assay

  • Species reactivity

    Reacts with: Human

  • Product overview

    Human Cleaved PARP1 SimpleStep ELISA® kit is a single-wash 90 min sandwich ELISA designed for the quantitative measurement of cleaved PARP1 protein in human cell extracts. Quantitate human cleaved PARP1 with 5.8 µg/ml sensitivity.

    SimpleStep ELISA® technology employs capture antibodies conjugated to an affinity tag that is recognized by the monoclonal antibody used to coat our SimpleStep ELISA® plates. This approach to sandwich ELISA allows the formation of the antibody-analyte sandwich complex in a single step, significantly reducing assay time. See the SimpleStep ELISA® protocol summary in the image section for further details. Our SimpleStep ELISA® technology provides several benefits:

            -Single-wash protocol reduces assay time to 90 minutes or less
            -High sensitivity, specificity and reproducibility from superior antibodies
            -Fully validated in biological samples
            -96-wells plate breakable into 12 x 8 wells strips

    A 384-well SimpleStep ELISA® microplate (ab203359) is available to use as an alternative to the 96-well microplate provided with SimpeStep ELISA® kits.

  • Notes

    PARP1 is a 113 kDa nuclear DNA-repair enzyme that transfers ADP-ribose units from NAD+ to variety of nuclear proteins including topoisomerases, histones and PARP1 itself. Via poly ADP ribosylation, PARP1 is responsible for regulation of cellular homeostasis including cellular repair, transcription and replication of DNA, cytoskeletal organization and protein degradation. In response to DNA damage, PARP1 activity is increased upon binding to DNA strand nicks and breaks. Excessive DNA damage leads to generation of large branched ADP-ribose polymers and activation of a unique cell death program.

    During apoptosis, PARP1 is cleaved by activated caspase-3 between Asp214 and Gly215, resulting in the formation of an N-terminal 24 kDa fragment containing most of the DNA binding domain and a C-terminal 89 kDa fragment containing the catalytic domain. The proteolysis of PARP1 through this cleavage renders the enzyme inactive and this further facilitates apoptotic cell death. Thus the presence of 89 kDa PARP1 fragment is considered to be a very reliable biomarker of apoptosis.

  • Platform

    Microplate

Properties

Associated products

Images

  • Other - Human Cleaved PARP1 ELISA Kit (ab174441)
    Other - Human Cleaved PARP1 ELISA Kit (ab174441)

    SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

     

  • Example of HeLa Staurosporine (ab120056) treated cell lysate titration.
    Example of HeLa Staurosporine (ab120056) treated cell lysate titration.
    Background-subtracted data values (mean +/- SD) are graphed.
  • HeLa and Jurkat cells were treated with 1 µM Staurosporine (STS) for 4 hours in complete cell culture media to induce apoptosis and cleaved PARP1 protein.
    HeLa and Jurkat cells were treated with 1 µM Staurosporine (STS) for 4 hours in complete cell culture media to induce apoptosis and cleaved PARP1 protein.

    Untreated and staurosporine (ab120056) treated HeLa and Jurkat lysates were prepared in 1X Cell Extraction Buffer PTR and tested using the Cleaved PARP1 SimpleStepELISA. Raw OD 450 nm values are shown for 500 µg/mL lysate loads.

  • Example of IC50 determination.
    Example of IC50 determination.

    HeLa cells were treated with a dose titration of Staurosporine for 4 hours in complete media. Cells were cultured and treated in a 96-well cell culture microtiter plate. Lysates were prepared by direct in-well lysis without media removal: 2X Cell Extraction Buffer PTR was added to an equal volume of media and then resulting lysate was used directly in the Cleaved PARP1 SimpleStepTM ELISA assay. Raw values for triplicate measurements are plotted. The calculated IC50 is 0.77 µM.

  • Demonstration of Cleaved PARP1 capture antibody specificity by western blot assay.
    Demonstration of Cleaved PARP1 capture antibody specificity by western blot assay.

    20 µg of HeLa extracts that were untreated or treated for 4 hours with 1 µM Staurosporine were analyzed by western blot. The GAPDH blot is included to show the relative loads of each lysate. In the HeLa cell line, Staurosporine treatment is required to detect cleaved PARP1 protein, as observed in the SimpleStep ELISA (Figure 2).

Protocols

References

This product has been referenced in:

  • Zhu J  et al. Nimotuzumab enhances the sensitivity of non-small cell lung cancer cells to tumor necrosis factor-a by inhibiting the nuclear factor-?B signaling pathway. Exp Ther Med 15:3345-3351 (2018). Read more (PubMed: 29545853) »
  • de Oliveira MR  et al. Pinocembrin Suppresses H2O2-Induced Mitochondrial Dysfunction by a Mechanism Dependent on the Nrf2/HO-1 Axis in SH-SY5Y Cells. Mol Neurobiol 55:989-1003 (2018). Read more (PubMed: 28084593) »
See all 2 Publications for this product

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