DNA DAMAGE IN HUMAN AND MOUSE SPERMATOZOA AFTER IN VITRO-IRRADIATION ASSESSED BY THE COMET ASSAY

摘要: The comet assay is widely employed as a method to measure DNA damage in wide variety of cell types following genotoxic insult. We have used this order characterise spermatozoa vitro irradiation with 137Cs gamma rays. In contrast somatic cells, the mammalian bound by protamine molecules allowing sixfold more highly compact structure and thus rendering conventional lysis protocols ineffective. Therefore, new uses an extensive step ensure effective removal DNA-associated proteins be scored reproducibly both murine human spermatozoa. Mouse collected from vas deferens at post-mortem or provided donors were irradiated doses γ-rays 0–100 Gy using source then processed for alkaline neutral assays. Under electrophoresis conditions, which permits measurement double-stranded breaks, linear increase amount measured was observed increasing radiation dose Similarly, conditions examine single-strand breaks alkali-labile sites, relationship also sperm but no such apparent subjected same treatments. Interestingly, unirradiated (both mouse) showed migration nucleus after assay. Since it unlikely that normal contains high numbers not detected assay, likely due presence alkali labile sites within these may related condensed spermatozoal DNA. large experiments produce measurable amounts reflects radioresistance compared cells differences packaging conformation. conclusion, work shows represents examining should evaluated use reproductive toxicity testing.