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Updated: Feb 26, 2026
Quantitation of γH2AX Foci in Tissue Samples
Published on: June 28, 2010
Express γ-H2AX Immunocytochemical Detection of DNA Damage
Nate Hopp1, Jodi Hagen1, Birte Aggeler1
1Bio-Techne, 614 McKinley Place NE, Minneapolis, MN, 55413, USA.
Abstract:
DNA can be damaged by many environmental factors including chemical agents and ionizing radiation which induce the formation of DNA double-stranded breaks (DSBs). If DSBs are not repaired in a timely fashion this may cause the disruption of genome integrity, which can result in cancer development. Typically, DSBs are followed by phosphorylation of histone protein H2AX, a member of the H2A family. Immunocytochemical detection of phosphorylated H2AX (e.g., γ-H2AX) appears to be a useful technique for assessing DNA damage. Such an assessment is easy to do by analyzing labeling for γ-H2AX under the microscope and does not require an expensive laboratory setup. Using HeLa cells treated with camptothecin as a model, we developed an easy-to-run protocol to analyze DSBs. Our protocol can be applied to testing the potency of different chemicals to induce DSBs in different types of cells and requires around 2 h to complete.
Insights
Environmental factors cause DNA double-stranded breaks (DSBs), potentially leading to cancer. A new, rapid immunocytochemical method using γ-H2AX detection can assess this DNA damage in cells.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Environmental factors like chemicals and radiation induce DNA double-stranded breaks (DSBs).
- Unrepaired DSBs can disrupt genome integrity, increasing cancer risk.
- Phosphorylation of histone H2AX (forming γ-H2AX) is a marker for DSBs.
Purpose of the Study:
- To develop a simple, rapid protocol for assessing DNA double-stranded breaks (DSBs).
- To utilize immunocytochemical detection of γ-H2AX for DNA damage assessment.
- To establish a model for testing chemical agents' potency in inducing DSBs.
Main Methods:
- Utilized HeLa cells treated with camptothecin as a model system.
- Employed immunocytochemical detection of phosphorylated H2AX (γ-H2AX).
- Developed a protocol requiring minimal laboratory equipment and approximately 2 hours.
Main Results:
- Successfully established an easy-to-run protocol for DSB analysis.
- Demonstrated the utility of γ-H2AX labeling for microscopic assessment of DNA damage.
- Validated the protocol's applicability across different cell types and chemical treatments.
Conclusions:
- The developed protocol provides a facile and efficient method for evaluating DNA damage induced by various agents.
- This technique is valuable for assessing the genotoxicity of chemicals and understanding DNA repair mechanisms.
- The method's simplicity and speed make it suitable for diverse research settings.
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