Urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine analysis by an improved ELISA: An inter-laboratory comparison study

0459609 - ÚEM 2017 RIV US eng J - Článek v odborném periodiku
Rössner ml., Pavel - Orhan, H. - Koppen, G. - Sakai, K. - Santella, R. M. - Ambrož, Antonín - Rössnerová, Andrea - Šrám, Radim - Ciganek, M. - Neca, J. - Arzuk, E. - Mutlu, N. - Cooke, M.S.
Urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine analysis by an improved ELISA: An inter-laboratory comparison study.
Free Radical Biology and Medicine. Roč. 95, č. 2016 (2016), s. 169-179. ISSN 0891-5849. E-ISSN 1873-4596
Grant CEP: GA ČR(CZ) GA13-13458S; GA MŠMT(CZ) LO1508; GA ČR(CZ) GA16-14631S
Institucionální podpora: RVO:68378041
Klíčová slova: 8-OxodG * ELISA * HPLC-MS/MS
Kód oboru RIV: DN - Vliv životního prostředí na zdraví
Impakt faktor: 5.606, rok: 2016

ELISA is commonly used for the detection of urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a marker of whole body oxidative stress. We performed an inter-laboratory comparison of 8-oxodG by ELISA using standardized experimental conditions, including: sample pre-treatment with solid-phase extraction (SPE), performing analysis using a commercial kit from a single manufacturer and strict temperature control during the assay. We compared the ELISA results with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and performed tentative identification of compounds that may contribute to the discrepancy between both methods. For all but one participating laboratory (Data 1) we observed consistent ELISA results lying mostly within 1SD of the mean 8-oxodG concentration. Mean 8-oxodG levels assessed by ELISA correlated with the data obtained by HPLC-MS/MS (R=0.679, p<0.001). The correlation improved when Data 1 were excluded from the analysis (R=0.749, p<0.001). We identified three outlying urine samples; one with an ELISA 8-oxodG concentration lower, and two with 8-oxodG levels higher, than those measured by HPLC-MS/MS. Omitting these samples further improved inter-methodology agreement (R=0.869, p<0.001). In the outliers with high 8-oxodG estimates various aromatic and heterocyclic compounds were tentatively identified using gas chromatography-mass spectrometry (GC-MS). Application of authentic standards revealed the presence of saccharides as putative interfering substances. In summary, assay standardization improved ELISA inter-laboratory agreement, although some variability is still observed. There are still compounds contributing to overestimation of 8-oxodG by ELISA, but only in some urine samples. Thus, despite significant improvement, ELISA still should not be considered a robust alternative to chromatographic techniques.
Trvalý link: http://hdl.handle.net/11104/0259797