{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,8]],"date-time":"2026-01-08T04:05:13Z","timestamp":1767845113325,"version":"3.49.0"},"reference-count":25,"publisher":"Wiley","issue":"5","license":[{"start":{"date-parts":[[2007,5,14]],"date-time":"2007-05-14T00:00:00Z","timestamp":1179100800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Reprod Domestic Animals"],"published-print":{"date-parts":[[2007,10]]},"abstract":"<jats:title>Contents<\/jats:title><jats:p>The main aim of this study was to compare the motility and functional integrity of bull spermatozoa after single and double freezing and thawing. The viability and morphological integrity of spermatozoa selected by PureSperm\u00ae density gradient centrifugation after cryopreservation of bovine semen in two commercial extenders (Experiment 1) and the function of bull spermatozoa before and after a second freezing and thawing assisted by PureSperm\u00ae selection (Experiment 2) were examined. On average, 35.8\u2003\u00b1\u200312.1% of sperm loaded onto the PureSperm\u00ae density gradient were recovered after centrifugation. In Experiment 1, post\u2010thaw motility and acrosome integrity were higher for spermatozoa frozen in Tris\u2010egg yolk extender than in AndroMed\u00ae, whether the assessments were made immediately after thawing [80.4\u2003\u00b1\u200312.7 vs 47.6\u2003\u00b1\u200319.0% motile and 78.8\u2003\u00b1\u20038.3 vs 50.1\u2003\u00b1\u200319.5% normal apical ridge (NAR), p\u2003&lt;\u20030.05] or after preparation on the gradient (83.3\u2003\u00b1\u20038.6 vs 69.4\u2003\u00b1\u200315.9% motile and 89.5\u2003\u00b1\u20037.2 vs 69.1\u2003\u00b1\u200311.4% NAR, p\u2003&lt;\u20030.05). For semen frozen in Tris\u2010egg yolk extender, selection on the PureSperm\u00ae gradient did not influence total motility but significantly improved the proportion of acrosome\u2010intact spermatozoa. After the gradient, both the total motility and percentage of normal acrosomes increased for spermatozoa frozen in AndroMed\u00ae (Minit\u00fcb Tiefenbach, Germany). In Experiment 2, there was no difference in sperm motility after the first and second freeze\u2010thawing (82.9\u2003\u00b1\u200312.7 vs 68.8\u2003\u00b1\u200318.7%). However, the proportion of acrosome\u2010intact spermatozoa was significantly improved by selection through the PureSperm\u00ae gradient, whether measured by phase contrast microscopy (78.9\u2003\u00b1\u20039.7 vs 90.4\u2003\u00b1\u20034.0% NAR, p\u2003&lt;\u20030.05) or flow cytometry (53.4\u2003\u00b1\u200311.7 vs 76.3\u2003\u00b1\u20036.0% viable acrosome\u2010intact spermatozoa, p\u2003&lt;\u20030.001). The improvement in the percentage of spermatozoa with normal acrosomes was maintained after resuspension in the cooling extender and cooling to 4\u00b0C (88.2\u2003\u00b1\u20036.2) and after re\u2010freezing and thawing (83.6\u2003\u00b1\u20036.56% NAR). However, flow cytometric assessment of the sperm membranes revealed a decline in the percentage of viable spermatozoa with intact membranes after the second freezing and thawing compared with after gradient centrifugation (76.3\u2003\u00b1\u20036.0% vs 46.6\u2003\u00b1\u20036.6%, p\u2003&lt;\u20030.001) to levels equivalent to those obtained after the first round of freeze\u2010thawing (53.4\u2003\u00b1\u200311.7% viable acrosome\u2010intact spermatozoa). Sperm movement characteristics assessed by computer\u2010assisted analysis were unaffected in the population selected on the PureSperm\u00ae gradients but declined after cooling of the selected and extended spermatozoa to 4\u00b0C. There was no further change in these kinematic measurements after the cooled spermatozoa had undergone the second round of freeze\u2010thawing. These results demonstrate that bull semen can be frozen and thawed, followed by a second freeze\u2010thawing cycle of a population of spermatozoa selected by PureSperm\u00ae, with retained motility and functional integrity. This points to the possibility of using double frozen spermatozoa in bovine artificial insemination programmes and to the potential benefits of PureSperm\u00ae density gradient centrifugation for the application of cryopreserved bull spermatozoa to other biotechnological procedures such as flow cytometric sex sorting followed by re\u2010freezing and thawing.<\/jats:p>","DOI":"10.1111\/j.1439-0531.2006.00811.x","type":"journal-article","created":{"date-parts":[[2007,5,14]],"date-time":"2007-05-14T09:40:08Z","timestamp":1179135608000},"page":"489-494","source":"Crossref","is-referenced-by-count":43,"title":["Retained Functional Integrity of Bull Spermatozoa after Double Freezing and Thawing Using PureSperm\u00ae Density Gradient Centrifugation"],"prefix":"10.1111","volume":"42","author":[{"given":"WMC","family":"Maxwell","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"I","family":"Parrilla","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"I","family":"Caballero","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"E","family":"Garcia","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"J","family":"Roca","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"EA","family":"Martinez","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"JM","family":"Vazquez","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"D","family":"Rath","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2007,5,14]]},"reference":[{"key":"e_1_2_6_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/0378-4320(93)90003-A"},{"key":"e_1_2_6_3_1","doi-asserted-by":"publisher","DOI":"10.1051\/rnd:2002044"},{"key":"e_1_2_6_4_1","doi-asserted-by":"publisher","DOI":"10.1093\/humrep\/13.11.3139"},{"key":"e_1_2_6_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.theriogenology.2006.08.005"},{"key":"e_1_2_6_6_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1600-0684.2005.00095.x"},{"key":"e_1_2_6_7_1","doi-asserted-by":"publisher","DOI":"10.1530\/rep.1.00049"},{"key":"e_1_2_6_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.theriogenology.2003.12.030"},{"key":"e_1_2_6_9_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.theriogenology.2004.04.002"},{"key":"e_1_2_6_10_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1439-0531.2006.00730.x"},{"key":"e_1_2_6_11_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1439-0531.2006.00729.x"},{"key":"e_1_2_6_12_1","doi-asserted-by":"publisher","DOI":"10.1016\/0011-2240(82)90121-3"},{"key":"e_1_2_6_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.anireprosci.2004.04.013"},{"key":"e_1_2_6_14_1","doi-asserted-by":"publisher","DOI":"10.1071\/RDv16n1Ab109"},{"key":"e_1_2_6_15_1","doi-asserted-by":"publisher","DOI":"10.1023\/B:JARG.0000040237.47026.0f"},{"key":"e_1_2_6_16_1","doi-asserted-by":"publisher","DOI":"10.1002\/j.1939-4640.2004.tb03157.x"},{"key":"e_1_2_6_17_1","doi-asserted-by":"publisher","DOI":"10.1095\/biolreprod.102.011445"},{"key":"e_1_2_6_18_1","doi-asserted-by":"publisher","DOI":"10.1071\/RD03065"},{"key":"e_1_2_6_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/0093-691X(74)90008-9"},{"key":"e_1_2_6_20_1","doi-asserted-by":"publisher","DOI":"10.1071\/R96081"},{"key":"e_1_2_6_21_1","doi-asserted-by":"publisher","DOI":"10.1093\/humrep\/15.5.1112"},{"key":"e_1_2_6_22_1","doi-asserted-by":"publisher","DOI":"10.3168\/jds.S0022-0302(72)85544-9"},{"key":"e_1_2_6_23_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.1439-0531.2002.00350.x"},{"key":"e_1_2_6_24_1","doi-asserted-by":"publisher","DOI":"10.1093\/humrep\/15.4.857"},{"key":"e_1_2_6_25_1","unstructured":"SteinbachJ 1963:Investigation on media and bovine methods for freezing bovine semen PhD thesis.Cornell University NY."},{"key":"e_1_2_6_26_1","first-page":"378","volume-title":"Reprod Dom Anim","author":"Underwood SL","year":"2006"}],"container-title":["Reproduction in Domestic Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1111%2Fj.1439-0531.2006.00811.x","content-type":"unspecified","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/j.1439-0531.2006.00811.x","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,10,14]],"date-time":"2023-10-14T16:53:36Z","timestamp":1697302416000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1111\/j.1439-0531.2006.00811.x"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2007,5,14]]},"references-count":25,"journal-issue":{"issue":"5","published-print":{"date-parts":[[2007,10]]}},"alternative-id":["10.1111\/j.1439-0531.2006.00811.x"],"URL":"https:\/\/doi.org\/10.1111\/j.1439-0531.2006.00811.x","archive":["Portico"],"relation":{},"ISSN":["0936-6768","1439-0531"],"issn-type":[{"value":"0936-6768","type":"print"},{"value":"1439-0531","type":"electronic"}],"subject":[],"published":{"date-parts":[[2007,5,14]]}}}