Evaluation of enzyme-linked immunosorbent assay and liquid chromatography-tandem mass spectrometry methodology for the analysis of 8-oxo-7,8-dihydro-2'-deoxyguanosine in saliva and urine.
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AuthorsCooke, Marcus S.
Hall, Georgina K.
Duarte, Tiago L.
Farmer, Peter B.
Evans, Mark D.
MetadataShow full item record
AbstractWhile ELISA is a frequently used means of assessing 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxodG) in biological fluids, differences in baseline urinary 8-oxodG levels, compared to chromatographic techniques, have raised questions regarding the specificity of immunoassays. Recently, ELISA of salivary 8-oxodG has been used to report on periodontal disease. We compared salivary 8-oxodG levels, determined by two commercial ELISA kits, to liquid chromatography-tandem mass spectrometry (LC-MS/MS) with prior purification using solid-phase extraction. While values were obtained with both ELISA kits, salivary 8-oxodG values were below or around the limit of detection of our LC-MS/MS assay. As the limit of detection for the LC-MS/MS procedure is much lower than ELISA, we concluded that the assessment of salivary 8-oxodG by ELISA is not accurate. In contrast to previous studies, ELISA levels of urinary 8-oxodG (1.67 +/- 0.53 pmol/mumol creatinine) were within the range reported previously only for chromatographic assays, although still significantly different than LC-MS/MS (0.41 +/- 0.39 pmol/mumol creatinine; p = 0.002). Furthermore, no correlation with LC-MS/MS was seen. These results question the ability of ELISA approaches, at present, to specifically determine absolute levels of 8-oxodG in saliva and urine. Ongoing investigation in our laboratories aims to identify the basis of the discrepancy between ELISA and LC-MS/MS.
CitationFree Radic. Biol. Med. 2006, 41 (12):1829-1836
JournalFree Radical Biology & Medicine
SponsorsThe authors gratefully acknowledge financial support of their laboratories from the Food Standards Agency; Medical Research Council; the authors of this paper are partners of ECNIS (Environment Cancer Risk, Nutrition and Individual Susceptibility), a network of excellence operating within the European Union 6th Framework Program, Priority 5: “Food Quality and Safety” (Contract No. 513943) The Biochemical Society. Creatinine determination by the Department of Chemical Pathology, Leicester Royal Infirmary, University Hospitals of Leicester NHS Trust, is also gratefully acknowledged. Statistical advice from Dr. Nick Taub, University of Leicester, is much appreciated.
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