{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,21]],"date-time":"2026-03-21T01:11:48Z","timestamp":1774055508539,"version":"3.50.1"},"reference-count":32,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2021,7,1]],"date-time":"2021-07-01T00:00:00Z","timestamp":1625097600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Punjab Educational Endowment Fund (PEEF)","award":["(PEEF\/SSMS\/18\/222)"],"award-info":[{"award-number":["(PEEF\/SSMS\/18\/222)"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>Sperm DNA fragmentation compromises fertilization and early embryo development. Since spermatozoa lack the machinery to repair DNA damage, to improve the likelihood of establishing a healthy pregnancy, it is preferable to process ejaculates of stallions with a high sperm DNA fragmentation index (DFI) before artificial insemination or intracytoplasmic sperm injection. The aim of this study was to examine a modified flotation density gradient centrifugation (DGC) technique in which semen was diluted with a colloid solution (Opti-prepTM) to increase its density prior to layering between colloid layers of lower and higher density. The optimal Opti-prepTM solution (20\u201360%) for use as the bottom\/cushion layer was first determined, followed by a comparison between a modified sedimentation DGC and the modified flotation DGC technique, using different Opti-prepTM solutions (20%, 25% and 30%) as the top layer. Finally, the most efficient DGC technique was selected to process ejaculates from Friesian stallions (n = 3) with high sperm DFI (&gt;20%). The optimal Opti-prepTM solution for the cushion layer was 40%. The modified sedimentation technique resulted in two different sperm populations, whereas the modified flotation technique yielded three populations. Among the variants tested, the modified flotation DGC using 20% Opti-prepTM as the top layer yielded the best results; the average sperm recovery was 57%; the DFI decreased significantly (from 12% to 4%) and the other sperm quality parameters, including progressive and total motility, percentages of spermatozoa with normal morphology and viable spermatozoa with an intact acrosome, all increased (p &lt; 0.05). In Friesian stallions with high sperm DFI, the modified flotation DGC markedly decreased the DFI (from 31% to 5%) and significantly improved the other semen quality parameters, although sperm recovery was low (approximately 20%). In conclusion, stallion sperm DFI and other sperm quality parameters can be markedly improved using a modified flotation DGC technique employing a 40% Opti-prepTM cushion and a 20% top layer.<\/jats:p>","DOI":"10.3390\/ani11071973","type":"journal-article","created":{"date-parts":[[2021,7,1]],"date-time":"2021-07-01T05:00:10Z","timestamp":1625115610000},"page":"1973","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["A Modified Flotation Density Gradient Centrifugation Technique Improves the Semen Quality of Stallions with a High DNA Fragmentation Index"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4858-1193","authenticated-orcid":false,"given":"Muhammad","family":"Umair","sequence":"first","affiliation":[{"name":"Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584 CM Utrecht, The Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4064-7792","authenticated-orcid":false,"given":"Heiko","family":"Henning","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584 CM Utrecht, The Netherlands"},{"name":"Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, H\u00f6ltystra\u00dfe 10, D-31535 Neustadt, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5321-8095","authenticated-orcid":false,"given":"Tom A. E.","family":"Stout","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584 CM Utrecht, The Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Anthony","family":"Claes","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584 CM Utrecht, The Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,7,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"33","DOI":"10.1007\/s00441-005-1097-5","article-title":"DNA damage to spermatozoa has impacts on fertilization and pregnancy","volume":"322","author":"Lewis","year":"2005","journal-title":"Cell Tissue Res."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"56","DOI":"10.1016\/j.anireprosci.2016.01.017","article-title":"The sperm chromatin structure assay (SCSA\u00ae) and other sperm DNA fragmentation tests for evaluation of sperm nuclear DNA integrity as related to fertility","volume":"169","author":"Evenson","year":"2016","journal-title":"Anim. 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