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Comparison between High-Performance Liquid Chromatography and Enzyme-linked Immunosorbent Assay for the Determination of 8-Hydroxy-2'-deoxyguanosine in Human Urine¹

Institute for Environmental Sciences [K. S.], and Department of Food and Nutritional Sciences, Graduate School of Nutritional and Environmental Sciences [N. Y., N. K.], University of Shizuoka, Shizuoka 422-8526; Department of Environmental Oncology, University of Occupational and Environmental Health, Kitakyushu 807-8555 [H. K.]; and Department of Pathology and Biology of Diseases, Graduate School of Medicine, Kyoto University, Kyoto 606-8501 [S. T.], Japan

8-Hydroxy-2'-deoxyguanosine (8-OH-dG), which has been regarded as a potential marker of oxidative DNA damage induced by reactive oxygen species, was measured in human urine by a commercial ELISA using a monoclonal antibody N45.1 and by high-performance liquid chromatography (HPLC) coupled to an electrochemical detector (HPLC-ECD) to evaluate whether the ELISA system is applicable to human monitoring studies. The urine samples were collected from 120 healthy men ages 18–58 in a steel-manufacturing company. A good correlation (r = 0.833; P < 0.0001) was observed between the two methods on HPLC-purified 8-OH-dG fractions from 23 urine samples. The mean value (±SD) of 8-OH-dG (µg/g creatinine) was 5.47 (±2.97) by HPLC-ECD assay and 5.50 (±2.36) by ELISA. However, the correlation (r) between the two methods on 120 original urine samples was 0.460 [P < 0.001; mean value (±SD) of 8-OH-dG (µg/g creatinine) was 4.46 (±2.03) by the HPLC assay and 9.33 (±3.23) by ELISA]. ELISA estimates were about 2-fold higher than the HPLC estimates on original urine. For an unknown reason, 10% of the urine samples showed more than a 4-fold increase in value by ELISA. It is suggested that the ELISA system is applicable for comparative human monitoring studies. Prepurification of ‘samples is required to determine the absolute value of 8-OH-dG in individual urine samples by ELISA.

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