Measurement of oxidatively generated base damage to nucleic acids in cells: facts and artifacts View Full Text


Ontology type: schema:ScholarlyArticle     


Article Info

DATE

2012-12

AUTHORS

Jean Cadet, Thierry Douki, Jean-Luc Ravanat, J. Richard Wagner

ABSTRACT

The search for DNA biomarkers of oxidative stress has been hampered for several decades by the lack of relevant information on base oxidation products and the challenging issue of measuring low amounts of lesions, typically a few modifications within the range 106–108 normal bases. In addition and this was ignored for a long time, there is a risk of artifactual oxidation of overwhelming nucleobases during DNA extraction and subsequent workup that has led to overestimation of some base damage up to 2–3 orders of magnitude. The main aim of the survey is to critically review the available methods that have been developed for measuring oxidatively generated base damage in nuclear and mitochondrial DNA. Among the chromatographic methods, high-performance liquid chromatography associated with tandem mass spectrometry (HPLC–MS/MS) is the most accurate and versatile approach whereas HPLC–electrochemical detection (ECD) is restricted to electrochemically active modifications. These methods allow measuring several single oxidized pyrimidine and purine bases, tandem base lesions and interstrand DNA cross-links in nuclear DNA. As complementary analytical tools, enzymatic methods that associate DNA repair enzymes with either the alkaline comet assay or the alkaline elution technique are suitable for assessing low variations in the level of different classes of oxidatively generated DNA lesions. Most of the immunoassays suffer from a lack of specificity due to the occurrence of cross-reactivity with overwhelming normal bases. One major exception concerns the immunodetection of 5-hydroxymethylcytosine, produced in a relatively high yield as an epigenetic DNA modification. HPLC–MS/MS is now recognized as the gold standard for measuring oxidized bases and nucleosides in human fluids such as urine, saliva, and plasma. More... »

PAGES

55-74

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  • Identifiers

    URI

    http://scigraph.springernature.com/pub.10.1007/s12566-012-0029-6

    DOI

    http://dx.doi.org/10.1007/s12566-012-0029-6

    DIMENSIONS

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