Analysis of urinary 8-oxo-7,8-dihydro-purine-2'-deoxyribonucleosides by LC-MS/MS and improved ELISA.
Non-invasive monitoring of oxidative stress is highly desirable. Urinary 7,8-8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) is a biologically relevant and convenient analytical target. However, immunoassays can over-estimate levels of urinary 8-oxodG. Measurement of more than one DNA oxidation product in urine would be advantageous in terms of mechanistic information. Urines samples were analysed for 8-oxodG by solid-phase extraction/LC-MS/MS and ELISA. The solid-phase extraction/LC-MS/MS assay was also applied to the analysis of urinary 7,8-dihydro-8-oxo-2'-deoxyadenosine (8-oxodA). Concurring with previous reports, urinary 8-oxodG measured by ELISA was significantly higher than levels measured by LC-MS/MS. However, apparent improvement in the specificity of the commercially available Japanese Institute for the Control of Ageing (JaICA) ELISA brought mean LC-MS/MS and ELISA measurements of urinary 8-oxodG into agreement. Urinary 8-oxodA was undetectable in all urines, despite efficient recovery by solid phase extraction. Exploitation of the advantages of ELISA may be enhanced by a simple modification to the assay procedure, although chromatographic techniques still remain the 'gold standard' techniques for analysis of urinary 8-oxodG. Urinary 8-oxodA is either not present or below the limit of detection of the instrumentation.
Evans
Mark D
MD
Department of Cancer, Radiation and Oxidative Stress Section, Studies and Molecular Medicine, University of Leicester, Leicester Royal Infirmary, University Hospitals of Leicester, Leicester, UK. mde2@le.ac.uk
Singh
Rajinder
R
Mistry
Vilas
V
Sandhu
Karendeep
K
Farmer
Peter B
PB
Cooke
Marcus S
MS
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000328:Adult; D015415:Biomarkers / Q000652:urine; D002853:Chromatography, Liquid; D004249:DNA Damage; D003839:Deoxyadenosines / Q000652:urine*; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000652:urine; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D008297:Male; D018384:Oxidative Stress; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry; D055815:Young Adult
D015415:Biomarkers; D003839:Deoxyadenosines; C078251:2'-deoxy-7,8-dihydro-8-oxoadenosine; D000080242:8-Hydroxy-2'-Deoxyguanosine; D003849:Deoxyguanosine