Development and application of a novel immunoassay for measuring oxidative DNA damage in the environment.
We developed a facile, cost-effective competitive binding assay for the analysis of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) in DNA, using a polyclonal rabbit antiserum raised against an 8-oxodGuo hapten coupled to bovine serum albumin and radiolabeled synthetic ligand containing multiple 8-oxodGuo residues. This radioimmunoassay (RIA) displays a high affinity for 8-oxodGuo in DNA, with a detection limit of approximately 1 adduct in 10(5) bases of DNA. 8-oxodGuo standards for RIA were quantified by high-performance liquid chromatography and electrochemical detection in DNA diluted in methylene blue and exposed to visible light. As an initial application we quantified 8-oxodGuo in dosimeters deployed at increasing depths in the Southern Ocean during the austral spring of the 1998 field season or at the surface at Palmer Station, Antarctica, throughout the 1999 field season. Cyclobutane pyrimidine dimers (CPD) were quantified using an established RIA. We found that the frequency of both photoproducts decreased with depth. However, CPD induction was attenuated at a faster rate than 8-oxodGuo, correlating with the differential attenuation of solar ultraviolet wavelengths in the water column. CPD induction was closely related with ultraviolet-B radiation (UVB) attenuation, whereas the lower attenuation of 8-oxodGuo suggests that oxidative damage is more closely related to ultraviolet-A radiation (UVA) irradiance. The ratio of 8-oxodGuo: CPD was also found to covary with changes in stratospheric ozone concentrations at Palmer Station. These data demonstrate the usefulness of these assays for environmental photobiology and the potential for their use in studying the relative impacts of UVB versus UVA, including ozone depletion events.
Mitchell
David L
DL
Department of Carcinogenesis, The University of Texas MD Anderson Cancer Center, Science Park/Research Division, Smithville 78957, USA. dmitchell@sprd1.mdacc.tmc.edu
Meador
Jarah
J
Paniker
Laxmi
L
Gasparutto
Didier
D
Jeffrey
Wade H
WH
Cadet
Jean
J
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D004249:DNA Damage; D004563:Electrochemistry; D004784:Environmental Monitoring; D006801:Humans; D018384:Oxidative Stress; D011863:Radioimmunoassay / Q000379:methods*; D013472:Sunlight