An electrochemiluminescence biosensor for 8-oxo-7,8-dihydro-2 '-deoxyguanosine quantification and DNA repair enzyme activity analysis using a novel bifunctional probe

摘要

A new electrochemiluminescence (ECL) sensor was developed for 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) quantification and Escherichia coil formamidopyrimidine-DNA glycosylase (FPG) activity assay. The sensor employed a novel spermine conjugated ruthenium tris-(bipyridine) derivative (spermine-Ru) which binds specifically with 8-oxodGuo through a one-step reaction and also acts as an ECL signal reporter. In the sensor, an 8-oxodGuo-containing ds-DNA film was first immobilized on a gold electrode by self-assembly. The DNA film was then incubated with spermine-Ru under oxidative condition for 8-oxodGuo labeling. The ECL intensity was found to correlate with the amount of 8-oxodGuo on the surface and the detection limit was estimated to be about 1 lesion in 500 DNA bases. Addition of FPG resulted in some loss of the signal due to the excision of 8-oxodGuo by the enzyme. An inverse relationship between ECL intensity and FPG concentration was observed in a range from 0 to 4.0 U/mu L, demonstrating that this sensor could be used for FPG activity assay. A number of metal ions were screened by the sensor for their inhibition effect on FPG activity. Among them, Hg2+ and methyl Hg(II) shown very potent inhibition, with IC50 values of 4.04 mu M and 4.34 nM respectively. The result may suggest that interference on the DNA repair system could be another mechanism for the high toxicity of MeHg. (C) 2015 Elsevier B.V. All rights reserved.