Effects of semen storage and separation techniques on sperm DNA fragmentation.
Abstract: To determine the effect of semen storage and separation techniques on sperm DNA fragmentation. Methods: Controlled clinical study. Methods: An assisted reproductive technology laboratory. Methods: Thirty normoozospermic semen samples obtained from patients undergoing infertility evaluation. Methods: One aliquot from each sample was immediately prepared (control) for the sperm chromatin dispersion assay (SCD). Aliquots used to assess storage techniques were treated in the following ways: snap frozen by liquid nitrogen immersion, slow frozen with Tris-yolk buffer and glycerol, kept on ice for 24 hours or maintained at room temperature for 4 and 24 hours. Aliquots used to assess separation techniques were processed by the following methods: washed and centrifuged in media, swim-up from washed sperm pellet, density gradient separation, density gradient followed by swim-up. DNA integrity was then measured by SCD. Methods: DNA fragmentation as measured by SCD. Results: There was no significant difference in fragmentation among the snap frozen, slow frozen, and wet-ice groups. Compared to other storage methods short-term storage at room temperature did not impact DNA fragmentation yet 24 hours storage significantly increased fragmentation. Swim-up, density gradient and density gradient/swim-up had significantly reduced DNA fragmentation levels compared with washed semen. Postincubation, density gradient/swim-up showed the lowest fragmentation levels. Conclusions: The effect of sperm processing methods on DNA fragmentation should be considered when selecting storage or separation techniques for clinical use.
Copyright © 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.
Publication Date: 2010-06-09 PubMed ID: 20542505DOI: 10.1016/j.fertnstert.2010.04.049Google Scholar: Lookup
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- Comparative Study
- Evaluation Study
- Journal Article
Summary
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This research investigates whether the ratio of X to Y chromosomes in horse sperm is affected by short-term storage or a swim-up procedure, concluding that neither method alters this ratio.
Objective of the research
- The aim of this study was to investigate whether the standard procedure of artificial insemination with fresh equine spermatozoa, which includes short-term storage and a swim-up procedure, affects the X/Y ratio in the sperm. This investigation is based on the fact that this procedure usually leads to a gradual loss of sperm viability.
Methodology
- The research uses a standard protocol for short-term storage of stallion semen, where it is stored for 0, 24 and 48 hours at 5°C, diluted in a commercial extender called EquiPro™.
- After each storage period, the motile fraction of sperm cells, or those cells that are capable of moving and therefore likely to fertilize an egg, are selected by a technique known as the swim-up method.
- The chromosomes in the sperm, particularly the X and Y chromosomes which determine the sex of offspring, are evaluated through fluorescence in situ hybridization (FISH). This method allows researchers to detect and localize specific DNA sequences on chromosomes.
Results
- The research found that the X/Y ratio in all sperm samples, both fresh and stored, did not differ from the expected 1:1 ratio, suggesting that neither short-term storage nor the swim-up method affect this ratio.
- The incidence of chromosomally abnormal sperm cells was also found to be insignificantly different in fresh and motile sperm samples.
Conclusion
- The study concludes that the two approaches being studied, sperm storage for up to 48 hours and the swim-up procedure, do not affect the X/Y ratio in the motile fraction of equine spermatozoa.
- This finding contradicts what has been described for human and cattle sperm, suggesting that there may be species-related differences in how sperm cells are affected by these procedures.
Cite This Article
APA
Jackson RE, Bormann CL, Hassun PA, Rocha AM, Motta EL, Serafini PC, Smith GD.
(2010).
Effects of semen storage and separation techniques on sperm DNA fragmentation.
Fertil Steril, 94(7), 2626-2630.
https://doi.org/10.1016/j.fertnstert.2010.04.049 Publication
Researcher Affiliations
- Department of Urology, University of Michigan, Ann Arbor, Michigan 48109, USA.
MeSH Terms
- Algorithms
- Cell Separation / methods
- Cytogenetic Analysis
- DNA / analysis
- DNA / metabolism
- DNA Fragmentation
- Humans
- Infertility, Male / genetics
- Male
- Reproductive Techniques, Assisted
- Semen Analysis
- Semen Preservation / adverse effects
- Semen Preservation / methods
- Sperm Retrieval / adverse effects
- Spermatozoa / metabolism
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