Development and validation of a new, sensitive immunochemical assay for O⁶-methylguanine in DNA and its application in a population study.

2.50
Hdl Handle:
http://hdl.handle.net/10146/196646
Title:
Development and validation of a new, sensitive immunochemical assay for O⁶-methylguanine in DNA and its application in a population study.
Authors:
Georgiadis, Panagiotis; Kaila, Stella; Makedonopoulou, Paraskevi; Fthenou, Eleni; Chatzi, Leda; Pletsa, Vasiliki; Kyrtopoulos, Soterios A.
Abstract:
Investigations of the presence of the precarcinogenic DNA adduct O⁶-methylguanine (O6-meG) in humans and its association with exposure or cancer risk have been hindered by the absence of analytic methods of adequate sensitivity and throughput. We report the development, validation, and application of an ELISA-type assay for O6-meG appropriate for large-scale population studies.; In the new analytic method, restriction enzymes are used to digest DNA to fragments of size expected to contain no more than one O6-meG residue. Anti-adduct antisera are used to transfer O6-meG-containing fragments to a solid surface, where they are detected using anti-ssDNA antisera, the high ratio of normal nucleotides to adducts providing a strong signal enhancement.; An assay with a limit of detection of 1.5 adducts/10⁹ nucleotides using 10 μg of DNA, a dynamic range of approximately two orders of magnitude and satisfactory precision and accuracy characteristics was established and validated. Analysis of samples from 120 subjects from the Rhea mother-child cohort in Crete led to the detection of O6-meG in 70% of maternal and 50% of cord blood buffy coat samples at mean levels of 0.65 and 0.38 adducts/10⁸ nucleotides, respectively.; The frequent observation of O6-meG in human DNA is compatible with dietary compounds (e.g. N-nitroso compounds or their precursors), or endogenous processes being responsible for the formation of this adduct.; The new assay opens the way for large-scale population studies of O6-meG as a biomarker of exposure or risk. The approach used in this assay can, in principle, be extended to any DNA adduct for which suitable antisera are available.
Citation:
Cancer Epidemiol. Biomarkers Prev. 2011, 20 (1):82-90
Journal:
Cancer Epidemiology, Biomarkers & Prevention
Issue Date:
Jan-2011
URI:
http://hdl.handle.net/10146/196646
DOI:
10.1158/1055-9965.EPI-10-0788
PubMed ID:
21081711
Additional Links:
http://cebp.aacrjournals.org/content/20/1/82.long
Type:
Article
Language:
en
ISSN:
1538-7755
Sponsors:
Research in this article was supported by contracts from the European Union (Network of Excellence ECNIS—"Environmental cancer risk, nutrition and individual susceptibility," contract number FOOD-CT-2005-513943;
Appears in Collections:
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Full metadata record

DC FieldValue Language
dc.contributor.authorGeorgiadis, Panagiotisen
dc.contributor.authorKaila, Stellaen
dc.contributor.authorMakedonopoulou, Paraskevien
dc.contributor.authorFthenou, Elenien
dc.contributor.authorChatzi, Ledaen
dc.contributor.authorPletsa, Vasilikien
dc.contributor.authorKyrtopoulos, Soterios A.en
dc.date.accessioned2011-12-09T12:47:06Z-
dc.date.available2011-12-09T12:47:06Z-
dc.date.issued2011-01-
dc.identifier.citationCancer Epidemiol. Biomarkers Prev. 2011, 20 (1):82-90en
dc.identifier.issn1538-7755-
dc.identifier.pmid21081711-
dc.identifier.doi10.1158/1055-9965.EPI-10-0788-
dc.identifier.urihttp://hdl.handle.net/10146/196646-
dc.description.abstractInvestigations of the presence of the precarcinogenic DNA adduct O⁶-methylguanine (O6-meG) in humans and its association with exposure or cancer risk have been hindered by the absence of analytic methods of adequate sensitivity and throughput. We report the development, validation, and application of an ELISA-type assay for O6-meG appropriate for large-scale population studies.en
dc.description.abstractIn the new analytic method, restriction enzymes are used to digest DNA to fragments of size expected to contain no more than one O6-meG residue. Anti-adduct antisera are used to transfer O6-meG-containing fragments to a solid surface, where they are detected using anti-ssDNA antisera, the high ratio of normal nucleotides to adducts providing a strong signal enhancement.en
dc.description.abstractAn assay with a limit of detection of 1.5 adducts/10⁹ nucleotides using 10 μg of DNA, a dynamic range of approximately two orders of magnitude and satisfactory precision and accuracy characteristics was established and validated. Analysis of samples from 120 subjects from the Rhea mother-child cohort in Crete led to the detection of O6-meG in 70% of maternal and 50% of cord blood buffy coat samples at mean levels of 0.65 and 0.38 adducts/10⁸ nucleotides, respectively.en
dc.description.abstractThe frequent observation of O6-meG in human DNA is compatible with dietary compounds (e.g. N-nitroso compounds or their precursors), or endogenous processes being responsible for the formation of this adduct.en
dc.description.abstractThe new assay opens the way for large-scale population studies of O6-meG as a biomarker of exposure or risk. The approach used in this assay can, in principle, be extended to any DNA adduct for which suitable antisera are available.en
dc.description.sponsorshipResearch in this article was supported by contracts from the European Union (Network of Excellence ECNIS—"Environmental cancer risk, nutrition and individual susceptibility," contract number FOOD-CT-2005-513943;en
dc.language.isoenen
dc.relation.urlhttp://cebp.aacrjournals.org/content/20/1/82.longen
dc.subjectDNA adducten
dc.subjectCancer risken
dc.subjectGuanineen
dc.subjectAnimalsen
dc.subjectHumansen
dc.subjectSensitivityen
dc.subjectDNAen
dc.subjectEnzyme-Linked Immunosorbent Assayen
dc.subject.meshAnimals-
dc.subject.meshBlood Buffy Coat-
dc.subject.meshDNA-
dc.subject.meshEnzyme-Linked Immunosorbent Assay-
dc.subject.meshFemale-
dc.subject.meshFetal Blood-
dc.subject.meshGuanine-
dc.subject.meshHela Cells-
dc.subject.meshHumans-
dc.subject.meshPregnancy-
dc.subject.meshRats-
dc.subject.meshRats, Sprague-Dawley-
dc.subject.meshReproducibility of Results-
dc.subject.meshSensitivity and Specificity-
dc.titleDevelopment and validation of a new, sensitive immunochemical assay for O⁶-methylguanine in DNA and its application in a population study.en
dc.typeArticleen
dc.identifier.journalCancer Epidemiology, Biomarkers & Preventionen
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