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EVALUATION OF DNA INTEGRITY USING TUNEL AND COMET ASSAY IN HUMAN SEMEN: IMMEDIATE- VERSUS DELAYED-FREEZING
Citation:
Young, K., L. Xun, S. A. Rothmann, S P. Darney, AND W. A. Robbins. EVALUATION OF DNA INTEGRITY USING TUNEL AND COMET ASSAY IN HUMAN SEMEN: IMMEDIATE- VERSUS DELAYED-FREEZING. Presented at American Society of Andrology, Phoenix, AZ, March 29-April 1, 2003.
Description:
EVALUATION OF DNA INTEGRITY USING TUNEL AND COMET ASSAY IN HUMAN SEMEN: IMMEDIATE- VERSUS DELAYED-FREEZING
K. Young,* L. Xun,* S. Rothmann,? S. Perreault, ? W. Robbins*
*University of California, Los Angeles, Los Angeles, California; ?Fertility Solutions Inc., Cleveland, Ohio; ?National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Research Triangle Park, North Carolina
Participation rates and geographic diversity in reproductive epidemiology studies would be greatly facilitated by home collection of ejaculated semen. To address concerns that home collection and mail return of semen would induce DNA damage, semen from 10 healthy men was collected. Part of each sample was snap frozen and the rest held at RT for 24-h (simulating overnight mail return) then snap frozen. DNA fragmentation was compared between the two groups using TUNEL and neutral comet assay. The TUNEL assay detected no significant difference between sperm fresh frozen (FF) and sperm frozen after 24-h (F24). Mean frequency and standard error per 2,000 cells scored was 136 ? 29 TUNEL positive cells in sperm FF versus 213 ? 28 TUNEL positive cells in sperm F24 (paired t-test; p = 0.07). The neutral comet assay detected a significant difference in double stranded DNA fragmentation. Mean comet tail length and standard error per 100 sperm cells scored was 174.6 ? 4.2 in sperm FF versus 146.1 ? 4.6 in sperm F24 (Wilcoxon signed rank test; p< 0.0001). Current findings indicate double stranded DNA damage is not induced in spermatozoa that remain in seminal plasma at RT for 24-h (similar to mailing conditions). The TUNEL assay may not have had enough power (75%) to detect a statistically significant difference between the two groups suggesting further study using larger sample sizes is necessary. This abstract does not reflect EPA policy; funded in part by EPA 9D-2295-NAEX.