QUANTITATIVE IMMUNOHISTOCHEMICAL DETERMINATION OF 8-HYDROXY-2'-DEOXYGUANOSINE BY A MONOCLONAL-ANTIBODY N45.1 - ITS APPLICATION TO FERRIC NITRILOTRIACETATE-INDUCED RENAL CARCINOGENESIS MODEL

The DNA base-modified product 8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the most commonly used markers for the evaluation of oxidative DNA damage. A monoclonal antibody specific for 8-OHdG (N45.1) was char acterized and applied in quantitative immunohistochemistry. N45.1 reco gnized both the modified base and deoxyribose structure of 8-OHdG and required a concentration two orders higher of 8-hydroxyguanosine as a competitor in the ELISA. In addition, N45.1 did not cross-react with t he original four deoxyribonucleosides, other DNA base-modified product s such as 8-hydroxy-2'-deoxy-adenosine and O-6-methyl-2'-deoxyguanosin e, or urine components such as uric acid, creatine, and creatinine. A ferric nitrilotriacetate-induced rat renal carcinogenesis model was us ed for the evaluation of quantitative immunohistochemistry. The 8-OHdG index of quantitative immunohistochemistry, as analyzed by NIH Image freeware, correlated reasonably well with the 8-OHdG amount determined by high-performance liquid chromatography with an electrochemical det ector-except for a difference in peak time, which could be attributed to the selection of target location. The present method has advantages over the high-performance liquid chromatography/electrochemical detec tor, gas chromatography/mass spectrometry, and P-32-postlabeling metho ds in that it allows localization of 8-OHdG to be specified without th e risk of artifactual production of 8-OHdG during the DNA extraction a nd hydrolytic processes.