Comet assay in human biomonitoring studies: reliability, validation, and applications.
The comet assay (single-cell gel electrophoresis), which measures DNA strand breaks at the level of single cells, is very easily applied to human lymphocytes, and therefore lends itself to human biomonitoring studies. For the examination of DNA base oxidation (a specific marker of oxidative damage), the assay is modified by including a stage at which the DNA is incubated with a suitable lesion-specific endonuclease. Here we report on the reliability and reproducibility of this approach, from the level of comparing results from duplicate gels prepared from the same sample of cells, up to an assessment of the natural intra- and interindividual variability in lymphocyte DNA damage measured in groups of normal, healthy human volunteers. We applied the assay in investigations of human disease and occupational exposure of factory workers.
Collins
A
A
Rowett Research Institute, Bucksburn, Aberdeen, UK. a.collins@rri.sari.ac.uk
Dusinská
M
M
Franklin
M
M
Somorovská
M
M
Petrovská
H
H
Duthie
S
S
Fillion
L
L
Panayiotidis
M
M
Raslová
K
K
Vaughan
N
N
D000704:Analysis of Variance; D000975:Antioxidants / Q000627:therapeutic use; D004249:DNA Damage; D004260:DNA Repair; D043382:DNA-Formamidopyrimidine Glycosylase; D043224:Deoxyribonuclease (Pyrimidine Dimer); D003922:Diabetes Mellitus, Type 1 / Q000235:genetics*; D004586:Electrophoresis / Q000379:methods*; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D004784:Environmental Monitoring / Q000379:methods*; D029968:Escherichia coli Proteins; D005821:Genetic Techniques; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007091:Image Processing, Computer-Assisted; D008214:Lymphocytes / Q000187:drug effects; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism; D016273:Occupational Exposure; D010084:Oxidation-Reduction; D015203:Reproducibility of Results
D000975:Antioxidants; D029968:Escherichia coli Proteins; D006861:Hydrogen Peroxide; D004706:Endodeoxyribonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase