Assay of excised oxidative DNA lesions: Isolation of 8-oxoguanine and its nucleoside derivatives from biological fluids with a monoclonal antibody column

Eun M I Park, Mark K. Shigenaga, Paolo Degan, Tommy S. Korn, Jeffrey W. Kitzler, Carol M. Wehr, Prema Kolachana, Bruch N. Ames

Research output: Contribution to journalArticle

Abstract

An immunoaffinity column is described that facilitates the analysis of oxidative damage products of DNA and RNA in urine, blood plasma, and medium isolated from cultures of Escherichia coli. In intact animals, lesions (adducts) excised from DNA are transported from the cell through the circulation and excreted in urine. In bacteria, DNA adducts are excreted directly into the medium. In either case, the adducts can be assayed as a measure of oxidative damage to DNA. A monoclonal antibody that recognizes 8-oxo-7,8-dihydro-2′-deoxyguanosine (oxo8dG; 8-hydroxy-2′-deoxyguanosine), a biomarker of oxidative damage to DNA, has been isolated, and its substrate binding properties have been characterized. The relative binding affinities of this monoclonal antibody for oxo8dG, unmodified nucleosides, or derivatives of Gua made it suitable for the preparation of immunoaffinity columns that greatly facilitate the isolation of oxo8dG, 8-oxo-7,8-dihydroguanine, and 8-oxo-7,8-dihydroguanosine from various biological fluids. Quantitative analysis of these adducts in wine of rats fed a nucleic acid-free diet and in the medium from cultures of E. coli suggests that oxo8-7,8-dihydroguanine is the principal repair product from oxo8dG in DNA of both eukaryotes and prokaryotes. The results support our previous estimate of about 105 oxidative lesions to DNA being formed and excised in an average rat cell per day.

Original languageEnglish
Pages (from-to)3375-3379
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume89
Issue number8
Publication statusPublished - 1992

Keywords

  • Cancer
  • Endogenous DNA adducts
  • Mutation
  • Oxygen radicals

ASJC Scopus subject areas

  • Genetics
  • General

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