Our genome experiences a moderate amount of DNA damage in our cells on a daily basis. This DNA damage can be in response to external environmental factors, or be a result of our internal metabolic processes going awry. While normal rates of DNA damage are not an immense threat to our cell processes, DNA damage in critical genes can lead to a variety of disease, including cancer and tumor formation. After induction of DNA damage (for example, in the form of double strand breaks), phosphorylation and recruitment of the H2AX protein occurs. This phosphorylation produces gamma H2AX, which is crucial for activating the DNA damage response (which in turn assembles DNA repair proteins at the mutation site) and creating checkpoint proteins to arrest the cell cycle. Gamma H2AX antibodies have been utilized to measure the toxic effect of a treatment or drug at the genotypic level, and are widely applied to a variety of research topics where DNA damage is at the forefront of disease pathology.
Immunohistochemistry-Paraffin: gamma H2AX [p Ser139] Antibody [NB100-384] - Sample: FFPE section of human ovarian carcinoma. Antibody: Affinity purified rabbit anti-gamma-H2AX used at a dilution of 1:5,000 (0.2ug/ml). Detection: DAB.
Jorge Mata-Gorrido et al used a gamma H2AX antibody in their study of neuronal accumulation of unrepaired DNA and how this affects aging and neurodegeneration. It has been previously reported that ATM regulates the DNA damage response in neurons, and that neurons are more at risk for DNA damage due to their high metabolic rate for energy production. In this study, a gamma H2AX antibody was used in immunofluorescence (IF) on ganglial neurons with x-ray radiation induced DNA damage. The gamma H2AX antibody was used primarily to track neuronal processing of repaired DNA over different time points post x-ray radiation. This experiment determined that DNA repair is a slow and long-term process in neurons. Next, a gamma H2AX antibody (alongside a 53BP1 antibody) was used to confirm that persistent DNA damaged foci were not a DNA free reservoir of protein components at the mutation site. Overall, it is clear that DNA damage is dynamic and dependent on the dose of environmental factor. In some foci the DNA damage is immediately repaired, and in others the DDR is prolonged.
Furthermore, Sofia Francia et al used a gamma H2AX antibody to establish that DNA damage response RNAs are required for the recruitment of DNA damage response factors. While it is already understood that DNA damage proteins are recruited to sites of DNA damage, Sofia Francia et al show that small non-coding RNAs are required to elicit this response. A gamma H2AX antibody was used in immunofluorescence staining of U2OS cells as a reference point for the level of DNA damage induced. It was concluded that DICER and DROSHA do not immediately sense DNA damage, but instead act as a secondary recruiter. Ultimately, it was predicted that gamma H2AX activation and formation comes before DICER and DROSHA generation. Whether these two pathways act together has still to be determined.
Novus Biologicals offers gamma H2AX reagents for your research needs including:
PMID: 20811597
PMID: 27102221
PMID: 26906421
