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However, the effect of SP on the sperm cryopreservation is dependent on the stallion and SP composition. The use of epididymal spermatozoa incubated in the presence of SP could help the identification of the components of SP that are able to confer protection upon the spermatozoa during freezing.<\/jats:p><\/jats:sec><jats:sec><jats:title>Objective<\/jats:title><jats:p>The aims of this study were (i) to identify SP components involved in the potential protection of epididymal spermatozoa during the freeze\u2010thawing process and (ii) to identify and evaluate the proteins likely related to sperm freezability, using two\u2010dimensional difference gel electrophoresis (2D\u2010DIGE).<\/jats:p><\/jats:sec><jats:sec><jats:title>Materials and Methods<\/jats:title><jats:p>Epididymal spermatozoa from 4 stallions were incubated with SP (80%, v\/v) or without SP (control) before freezing. Sperm parameters were evaluated after thawing (viability, chromatin condensation, acrosomal integrity, reactive oxygen species [ROS]) and SP composition: total antioxidant capacity (TAC), fatty acid composition, total protein concentration, and protein components by 2D\u2010DIGE.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>After thawing, the proportions of viable and acrosome\u2010intact spermatozoa were higher than control when SP from two stallions was used (F and O). The SP of all stallions reduced ROS production in comparison with the control. After analyzing the SP components, it was found that total protein concentration, TAC, polyunsaturated fatty acids (PUFA), and eight specific proteins identified by 2D\u2010DIGE were different between stallions.<\/jats:p><\/jats:sec><jats:sec><jats:title>Discussion<\/jats:title><jats:p>These studies allow the identification of SP components that could be involved in sperm protection or cryotolerance. Use of this information could help in the selection of stallions according to their semen freezing capacity.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusion<\/jats:title><jats:p>The composition of the SP probably contributes to semen cryotolerance capacity. Total protein, TAC, PUFA, and some proteins such as cysteine\u2010rich secreted protein 3 could be used as biomarkers for the selection for sperm cryotolerance.<\/jats:p><\/jats:sec>","DOI":"10.1111\/andr.12944","type":"journal-article","created":{"date-parts":[[2020,11,13]],"date-time":"2020-11-13T11:06:05Z","timestamp":1605265565000},"page":"728-743","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":10,"title":["Seminal plasma components from fertile stallions involved in the epididymal sperm freezability"],"prefix":"10.1111","volume":"9","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2419-1329","authenticated-orcid":false,"given":"Luis Alberto","family":"Vieira","sequence":"first","affiliation":[{"name":"Department of Physiology International Excellence Campus for Higher Education and Research (Campus Mare Nostrum) University of Murcia  Murcia Spain"},{"name":"Institute for Biomedical Research of Murcia IMIB\u2010Arrixaca 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