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The DNA Damage Assay is a simple, fast and accurate, 2-color fluorescent assay for the induction of double-stranded DNA (dsDNA) breaks. When cell cultures or animals are exposed to ionizing radiation or certain treatment compounds, double-stranded DNA breaks are created that rapidly result in the phosphorylation of the histone variant H2AX at serine 139. Because H2AX phosphorylation at serine 139 correlates very closely with each dsDNA break, phospho-H2AX is a sensitive marker for DNA damage and can be used to assess the effects and timing of DNA-damage inducing and DNA repair agents.
| Name | Format | Cat. No. | Price | |
|---|---|---|---|---|
| DNA Damage Assay (Fluorescent) | 2 x 96 rxns | 18030 | $405 |
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| DNA Damage Assay (Fluorescent) Manual |
| DNA Damage Assay (Fluorescent) Profile |
| Chromatin & Epigenetics Products Brochure |
The assay utilizes a highly specific DNA damage and apoptosis marker antibody, anti-phospho-Histone H2AX (Ser 139), which is detected by a fluorescent Chromeo™ 488-labeled secondary antibody. Cells are grown and treated in a 96-well plate, then fixed and incubated with the phospho-H2AX apoptosis marker antibody that is subsequently detected with the Chromeo 488 secondary. Lastly, cells are washed and stained with propidium iodide and the plate is read on a fluorescent plate reader.
The DNA Damage Assay is suited for high-throughput and high-content fluorescent scanning applications. Each kit contains reagents for two full 96-well plates and includes apoptosis marker antibody, fluorescent secondary antibody, propidium iodide and etoposide, which is a positive control compound that causes high levels of DNA damage (Figure 1).
Advantages
- Easy – Very little hands-on-time
- Accurate – Propidium iodide readout provides total cell count
- Specific – High-quality antibodies ensure no problems with background
Figure 1: Percent of HeLa cells positive for phospho-H2AX.
Levels of phospho-H2AX measured in untreated (control) and cells treated with etoposide.
Contents & Storage
2 x 96 rxns each of phospho-H2AX (S139) antibody, Chromeo™ 488 Goat anti-Rabbit IgG, propidium iodide, etoposide and RNase A. Reagents are to be stored at -20°C. All reagents are guaranteed stable for 6 months from date of receipt when stored properly.
