Electrochemiluminescent/voltammetric toxicity screening sensor using enzyme-generated DNA damage.
Simultaneous optical and voltammetric detection of bioactivated genotoxicity is reported for the first time employing ultrathin films of DNA, model metabolic enzymes, and electrochemiluminescence (ECL) generating metallopolymer [Ru(bpy)2PVP10]2+ on pyrolytic graphite (PG) electrodes. Cytochrome P450cam and myoglobin were used as model monoxygenase enzymes to mimic in vivo processes. Sensor film growth and component amounts were monitored using a quartz crystal microbalance (QCM). Subsequent to the enzyme reaction, DNA damage in the sensor films was measured simultaneously using a simple apparatus combining a standard voltammetry cell coupled with an optical fiber and photomultiplier tube. The model enzyme reaction converted styrene to styrene oxide, which reacts with DNA nucleobases. ECL and SWV signals increased with enzyme reaction time on the scale of several min, and provided relative enzyme turnover rates for DNA damage suitable for toxicity screening applications. Within 1 min, the sensor detects approximately 3 damaged bases per 10,000 DNA bases using this simultaneous detection.
So
Minjeong
M
Department of Chemistry, University of Connecticut, Storrs, CT 06269-3060, United States.
Hvastkovs
Eli G
EG
Schenkman
John B
JB
Rusling
James F
JF
D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D019475:Camphor 5-Monooxygenase / Q000737:chemistry / Q000378:metabolism; D004249:DNA Damage; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods*; D004566:Electrodes; D004798:Enzymes / Q000737:chemistry* / Q000378:metabolism; D006108:Graphite / Q000737:chemistry; D006861:Hydrogen Peroxide / Q000737:chemistry; D008163:Luminescent Measurements / Q000295:instrumentation / Q000379:methods*