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Measurement of superoxide dismutase, catalase and glutathione peroxidase in cultured cells and tissue

Nature Protocols volume 5, pages 5166 (2010) | Download Citation

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Abstract

Cells contain a large number of antioxidants to prevent or repair the damage caused by reactive oxygen species, as well as to regulate redox-sensitive signaling pathways. General protocols are described to measure the antioxidant enzyme activity of superoxide dismutase (SOD), catalase and glutathione peroxidase. The SODs convert superoxide radical into hydrogen peroxide and molecular oxygen, whereas the catalase and peroxidases convert hydrogen peroxide into water. In this way, two toxic species, superoxide radical and hydrogen peroxide, are converted to the harmless product water. Western blots, activity gels and activity assays are various methods used to determine protein and activity in both cells and tissue depending on the amount of protein required for each assay. Other techniques including immunohistochemistry and immunogold can further evaluate the levels of the various antioxidant enzymes in tissues and cells. In general, these assays require 24–48 h to complete.

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Acknowledgements

This work was supported by an NIH Grant CA137230 and a VA Merit Review Grant.

Author information

Author notes

    • Christine J Weydert
    •  & Joseph J Cullen

    These authors contributed equally to this work.

Affiliations

  1. Department of Molecular Physiology and Biophysics, The University of Iowa Carver College of Medicine, Iowa City, Iowa, USA.

    • Christine J Weydert
  2. Free Radical and Radiation Biology Program, Department of Radiation Oncology, The University of Iowa Carver College of Medicine, Iowa City, Iowa, USA.

    • Joseph J Cullen
  3. Holden Comprehensive Cancer Center, The University of Iowa Carver College of Medicine, Iowa City, Iowa, USA.

    • Christine J Weydert
    •  & Joseph J Cullen
  4. Department of Surgery, The University of Iowa Carver College of Medicine, Iowa City, Iowa, USA.

    • Joseph J Cullen
  5. VA Medical Center, Iowa City, Iowa, USA.

    • Joseph J Cullen

Authors

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Contributions

C.J.W. and J.J.C. discussed and commented on the paper at all stages.

Corresponding author

Correspondence to Joseph J Cullen.

Supplementary information

Word documents

  1. 1.

    Supplementary Method

    Method for fixing tissues and cultured cells for immunogold immunohistochemistry.

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DOI

https://doi.org/10.1038/nprot.2009.197

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