{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T00:54:03Z","timestamp":1792025643491,"version":"4.4.0"},"reference-count":80,"publisher":"MDPI AG","issue":"17","license":[{"start":{"date-parts":[[2020,9,3]],"date-time":"2020-09-03T00:00:00Z","timestamp":1599091200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Czech University of Life Sciences in Prague","award":["SV19-05-21230"],"award-info":[{"award-number":["SV19-05-21230"]}]},{"DOI":"10.13039\/501100001824","name":"Grantov\u00e1 Agentura \u010cesk\u00e9 Republiky","doi-asserted-by":"publisher","award":["GA-18-11275S"],"award-info":[{"award-number":["GA-18-11275S"]}],"id":[{"id":"10.13039\/501100001824","id-type":"DOI","asserted-by":"publisher"}]},{"name":"BIOCEV (CZ.1.05\/1.1.00\/02.0109) from ERDF and Institute of Biotechnology","award":["RVO:86652036"],"award-info":[{"award-number":["RVO:86652036"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Seminal plasma (SP) is the natural environment for spermatozoa and contains a number of components, especially proteins important for successful sperm maturation and fertilization. Nevertheless, in standard frozen stallion insemination doses production, SP is completely removed and is replaced by a semen extender. In the present study, we analyzed the effects of the selected seminal plasma protein groups that might play an important role in reducing the detrimental effects on spermatozoa during the cryopreservation process. SP proteins were separated according to their ability to bind to heparin into heparin-binding (Hep+) and heparin-non-binding (Hep\u2212) fractions. The addition of three concentrations\u2014125, 250, and 500 \u00b5g\/mL\u2014of each protein fraction was tested. After thawing, the following parameters were assessed: sperm motility (by CASA), plasma membrane integrity (PI staining), and acrosomal membrane integrity (PNA staining) using flow cytometry, and capacitation status (anti-phosphotyrosine antibody) using imaging-based flow cytometry. Our results showed that SP protein fractions had a significant effect on the kinematic parameters of spermatozoa and on a proportion of their subpopulations. The 125 \u00b5g\/mL of Hep+ protein fraction resulted in increased linearity (LIN) and straightness (STR), moreover, with the highest values of sperm velocities (VAP, VSL), also this group contained the highest proportion of the fast sperm subpopulation. In contrast, the highest percentage of slow subpopulation was in the groups with 500 \u00b5g\/mL of Hep+ fraction and 250 \u00b5g\/mL of Hep\u2212 fraction. Interestingly, acrosomal membrane integrity was also highest in the groups with Hep+ fraction in concentrations of 125 \u00b5g\/mL. Our results showed that the addition of protein fractions did not significantly affect the plasma membrane integrity and capacitation status of stallion spermatozoa. Moreover, our results confirmed that the effect of SP proteins on the sperm functionality is concentration-dependent, as has been reported for other species. Our study significantly contributes to the lack of studies dealing with possible use of specific stallion SP fractions in the complex puzzle of the improvement of cryopreservation protocols. It is clear that improvement in this field still needs more outputs from future studies, which should be focused on the effect of individual SP proteins on other sperm functional parameters with further implication on the success of artificial insemination in in vivo conditions.<\/jats:p>","DOI":"10.3390\/ijms21176415","type":"journal-article","created":{"date-parts":[[2020,9,3]],"date-time":"2020-09-03T08:40:26Z","timestamp":1599122426000},"page":"6415","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":28,"title":["Effect of Seminal Plasma Protein Fractions on Stallion Sperm Cryopreservation"],"prefix":"10.3390","volume":"21","author":[{"given":"Filipa","family":"Bubenickova","sequence":"first","affiliation":[{"name":"Department of Veterinary Sciences, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, 165 00 Prague, Czech Republic"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5090-4652","authenticated-orcid":false,"given":"Pavla","family":"Postlerova","sequence":"additional","affiliation":[{"name":"Department of Veterinary Sciences, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, 165 00 Prague, Czech Republic"},{"name":"Laboratory of Reproductive Biology, Institute of Biotechnology of the Czech Academy of Sciences, BIOCEV, 252 50 Vestec, Czech Republic"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ondrej","family":"Simonik","sequence":"additional","affiliation":[{"name":"Department of Veterinary Sciences, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, 165 00 Prague, Czech Republic"},{"name":"Laboratory of Reproductive Biology, Institute of Biotechnology of the Czech Academy of Sciences, BIOCEV, 252 50 Vestec, Czech Republic"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jitka","family":"Sirohi","sequence":"additional","affiliation":[{"name":"Department of Statistics, Faculty of Economics and Management, Czech University of Life Sciences Prague, 165 00 Prague, Czech Republic"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiri","family":"Sichtar","sequence":"additional","affiliation":[{"name":"Department of Veterinary Sciences, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, 165 00 Prague, Czech Republic"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,9,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"907","DOI":"10.1016\/S0093-691X(97)00319-1","article-title":"Equine frozen semen: Freezability and fertility field results","volume":"48","author":"Vidament","year":"1997","journal-title":"Theriogenology"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"895","DOI":"10.1016\/S0093-691X(98)00039-9","article-title":"Current methods for stallion semen cryopreservation: A survey","volume":"49","author":"Samper","year":"1998","journal-title":"Theriogenology"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1016\/S0378-4320(01)00156-7","article-title":"The equine frozen semen industry","volume":"68","author":"Loomis","year":"2001","journal-title":"Anim. Reprod. Sci."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1584","DOI":"10.1016\/j.theriogenology.2004.05.030","article-title":"Relationship of seminal plasma level and extender type to sperm motility and DNA integrity","volume":"63","author":"Love","year":"2005","journal-title":"Theriogenology"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1016\/j.anireprosci.2005.06.018","article-title":"The role of stallion seminal proteins in fertilisation","volume":"89","author":"Kirchhoff","year":"2005","journal-title":"Anim. Reprod. Sci."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"640","DOI":"10.1095\/biolreprod.103.016527","article-title":"Influence of porcine spermadhesins on the susceptibility of boar spermatozoa to high dilution","volume":"69","author":"Centurion","year":"2003","journal-title":"Biol. Reprod."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"700","DOI":"10.1002\/j.1939-4640.2000.tb02138.x","article-title":"Cryopreservation alters the levels of the bull sperm surface protein P25b","volume":"21","author":"Lessard","year":"2000","journal-title":"J. Androl."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1016\/j.cryobiol.2011.12.002","article-title":"Freezing injury: The special case of the sperm cell","volume":"64","author":"Morris","year":"2012","journal-title":"Cryobiology"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1016\/0011-2240(90)90009-S","article-title":"Cold-Induced ultrastructural-changes in bull and boar sperm plasma-membranes","volume":"27","author":"Deleeuw","year":"1990","journal-title":"Cryobiology"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1002\/j.1939-4640.2000.tb03268.x","article-title":"Semen cryopreservation in domestic animals: A damaging and capacitating phenomenon","volume":"21","author":"Bailey","year":"2000","journal-title":"J. Androl."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"724","DOI":"10.1111\/rda.12151","article-title":"Sequestration of PDC-109 Protein by Specific Antibodies and Egg Yolk Cryoprotects Bull Spermatozoa","volume":"48","author":"Srivastava","year":"2013","journal-title":"Reprod. Domest. Anim."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"545","DOI":"10.1530\/REP-09-0375","article-title":"Proteomic comparison of detergent-extracted sperm proteins from bulls with different fertility indexes","volume":"139","author":"Frenette","year":"2010","journal-title":"Reproduction"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"220","DOI":"10.1016\/j.anireprosci.2007.02.014","article-title":"Seminal plasma non-heparin binding proteins (NHBP) reduce the cryoinjury to buffalo cauda epididymal spermatozoa induced by heparin binding proteins (HBP)","volume":"104","author":"Kumar","year":"2008","journal-title":"Anim. Reprod. Sci."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"395","DOI":"10.1016\/j.anireprosci.2006.08.018","article-title":"Effect of egg yolk and seminal plasma heparin binding protein interaction on the freezability of buffalo cauda epididymal spermatozoa","volume":"99","author":"Singh","year":"2007","journal-title":"Anim. Reprod. Sci."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"124","DOI":"10.1016\/j.anireprosci.2005.07.010","article-title":"Effect of buffalo seminal plasma heparin binding protein (HBP) on freezability and in vitro fertility of buffalo cauda spermatozoa","volume":"93","author":"Harshan","year":"2006","journal-title":"Anim. Reprod. Sci."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1621","DOI":"10.1111\/rda.13571","article-title":"Extenders modify the seminal plasma ability to minimize freeze-thaw damage on ram sperm","volume":"54","author":"Cesari","year":"2019","journal-title":"Reprod. Domest. Anim."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"791","DOI":"10.1016\/S0093-691X(96)00237-3","article-title":"Seminal plasma affects membrane integrity and motility of equine spermatozoa after cryopreservation","volume":"46","author":"Aurich","year":"1996","journal-title":"Theriogenology"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1016\/j.cryobiol.2018.01.009","article-title":"Improved cryosurvival of stallion spermatozoa after colloid centrifugation is independent of the addition of seminal plasma","volume":"81","author":"Johannisson","year":"2018","journal-title":"Cryobiology"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2372","DOI":"10.1016\/j.theriogenology.2004.05.032","article-title":"Effect of seminal plasma on the cryopreservation of equine spermatozoa","volume":"63","author":"Moore","year":"2005","journal-title":"Theriogenology"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1016\/j.anireprosci.2008.05.084","article-title":"Effects of addition of seminal plasma on lifespan of frozen-thawed equine spermatozoa","volume":"107","author":"Arruda","year":"2008","journal-title":"Anim. Reprod. Sci."},{"key":"ref_21","first-page":"83","article-title":"Motility of frozen-thawed stallion epididymal sperm after the addition of homologous seminal plasma from stallions with different semen quality","volume":"49","author":"Neuhauser","year":"2014","journal-title":"Reprod. Domest. Anim."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Sichtar, J., Bubenickova, F., Sirohi, J., and Simonik, O. (2019). How to Increase Post-Thaw Semen Quality in Poor Freezing Stallions: Preliminary Results of the Promising Role of Seminal Plasma Added after Thawing. Animals, 9.","DOI":"10.3390\/ani9070414"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"682","DOI":"10.1111\/j.1439-0531.2010.01729.x","article-title":"Addition of Seminal Plasma to Post-thawing Equine Semen: What is the Effect on Sperm Cell Viability","volume":"46","author":"Zaffalon","year":"2011","journal-title":"Reprod. Domest. Anim."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1352","DOI":"10.1016\/j.theriogenology.2008.07.013","article-title":"Major proteins of boar seminal plasma as a tool for biotechnological preservation of spermatozoa","volume":"70","author":"Caballero","year":"2008","journal-title":"Theriogenology"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"193","DOI":"10.21836\/PEM20060214","article-title":"Components of stallion seminal plasma and their influence on spermatozoa","volume":"22","author":"Katila","year":"2006","journal-title":"Pferdeheilkunde"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1016\/j.anireprosci.2016.03.047","article-title":"Seminal plasma components differ between \u201cgood freezer\u201d and \u201cpoor freezer\u201d stallions","volume":"169","author":"Tsikis","year":"2016","journal-title":"Anim. Reprod. Sci."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"411","DOI":"10.1111\/j.1439-0531.1994.tb00588.x","article-title":"isolation and biochemical-characterization of stallion seminal-plasma proteins","volume":"29","author":"Calvete","year":"1994","journal-title":"Reprod. Domest. Anim."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"636","DOI":"10.1111\/j.1432-1033.1996.0636r.x","article-title":"Primary structure of stallion seminal plasma protein HSP-7, a zona-pellucida-binding protein of the spermadhesin family","volume":"242","author":"Reinert","year":"1996","journal-title":"Eur. J. Biochem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1016\/S0014-5793(97)01514-7","article-title":"Biochemical and conformational characterisation of HSP-3, a stallion seminal plasma protein of the cysteine-rich secretory protein (CRISP) family","volume":"420","author":"Magdaleno","year":"1997","journal-title":"FEBS Lett."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1002\/mrd.20187","article-title":"Structural and molecular characterization of equine sperm-binding fibronectin-II module proteins","volume":"70","author":"Schafer","year":"2005","journal-title":"Mol. Reprod. Dev."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"206","DOI":"10.1016\/S0167-4838(98)00122-8","article-title":"Equine CRISP-3: Primary structure and expression in the male genital tract","volume":"1387","author":"Schambony","year":"1998","journal-title":"Biochim. Biophys. Acta Protein Struct. Mol. Enzymol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"563","DOI":"10.1071\/RD17044","article-title":"Association of the cysteine-rich secretory protein-3 (CRISP-3) and some of its polymorphisms with the quality of cryopreserved stallion semen","volume":"30","author":"Usuga","year":"2018","journal-title":"Reprod. Fertil. Dev."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1247","DOI":"10.5713\/ajas.15.0586","article-title":"Seminal Plasma Heparin Binding Proteins Improve Semen Quality by Reducing Oxidative Stress during Cryopreservation of Cattle Bull Semen","volume":"29","author":"Patel","year":"2016","journal-title":"Asian Australas. J. Anim. Sci."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1016\/j.anireprosci.2005.03.014","article-title":"Fertility-associated proteins in Nelore bull sperm membranes","volume":"91","author":"Roncoletta","year":"2006","journal-title":"Anim. Reprod. Sci."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"956","DOI":"10.1016\/j.theriogenology.2015.05.035","article-title":"Two-dimensional polyacrylamide gel electrophoresis of Bali bull (Bos javanicus) seminal plasma proteins and their relationship with semen quality","volume":"84","author":"Sarsaifi","year":"2015","journal-title":"Theriogenology"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"956","DOI":"10.1016\/j.theriogenology.2010.04.025","article-title":"Biomarkers of in vivo fertility in sperm and seminal plasma of fertile stallions","volume":"74","author":"Novak","year":"2010","journal-title":"Theriogenology"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"765","DOI":"10.1016\/j.theriogenology.2011.04.010","article-title":"Two-dimensional polyacrylamide gel electrophoresis of equine seminal plasma proteins and their relation with semen freezability","volume":"76","author":"Jobim","year":"2011","journal-title":"Theriogenology"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1111\/rda.13309","article-title":"Relationship of cysteine-rich secretory protein-3 gene and protein with semen quality in stallions","volume":"54","author":"Restrepo","year":"2019","journal-title":"Reprod. Domest. Anim."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"157","DOI":"10.1095\/biolreprod.110.084491","article-title":"Equine CRISP3 Modulates Interaction Between Spermatozoa and Polymorphonuclear Neutrophils","volume":"85","author":"Doty","year":"2011","journal-title":"Biol. Reprod."},{"key":"ref_40","first-page":"69","article-title":"Effect of purified seminal plasma heparin binding proteins on in vitro acrosome reaction of frozen-thawed buffalo bull semen","volume":"93","author":"Singh","year":"2016","journal-title":"Indian Vet. J."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.anireprosci.2013.02.008","article-title":"Bovine seminal PDC-109 protein: An overview of biochemical and functional properties","volume":"138","author":"Srivastava","year":"2013","journal-title":"Anim. Reprod. Sci."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"955","DOI":"10.1016\/0093-691X(95)00045-A","article-title":"Effect of seminal extenders containing egg-yolk and glycerol on motion characteristics and fertility of stallion spermatozoa","volume":"43","author":"Bedford","year":"1995","journal-title":"Theriogenology"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"479","DOI":"10.1111\/j.1439-0531.2006.00810.x","article-title":"Practical techniques for bovine sperm simultaneous fluorimetric assessment of plasma, acrosomal and mitochondrial membranes","volume":"42","author":"Celeghini","year":"2007","journal-title":"Reprod. Domest. Anim."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"192","DOI":"10.1016\/j.anireprosci.2010.11.017","article-title":"Influence of seminal plasma on fresh and post-thaw parameters of stallion epididymal spermatozoa","volume":"123","author":"Heise","year":"2011","journal-title":"Anim. Reprod. Sci."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"De Lazari, F.L., Sontag, E.R., Schneider, A., Moura, A.A.A., Vasconcelos, F.R., Nagano, C.S., Mattos, R.C., Jobim, M.I.M., and Bustamante, I.C. (2019). Seminal plasma proteins and their relationship with sperm motility and morphology in boars. Andrologia, 51.","DOI":"10.1111\/and.13222"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"171","DOI":"10.56093\/ijans.v90i2.98769","article-title":"Lyophilized seminal plasma can improve stallion semen freezability","volume":"90","author":"Rojano","year":"2020","journal-title":"Indian J. Anim. Sci."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1973","DOI":"10.1016\/S0093-691X(02)01297-9","article-title":"Identification of sperm subpopulations with specific motility characteristics in stallion ejaculates","volume":"59","author":"Miro","year":"2003","journal-title":"Theriogenology"},{"key":"ref_48","first-page":"88","article-title":"Lipid Peroxidation, Assessed with BODIPY-C11, Increases after Cryopreservation of Stallion Spermatozoa, is Stallion-dependent and is Related to Apoptotic-like Changes","volume":"44","author":"Ferrusola","year":"2009","journal-title":"Reprod. Domest. Anim."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"783","DOI":"10.1016\/j.theriogenology.2010.11.034","article-title":"Statistical Series: Opportunities and challenges of sperm motility subpopulation analysis","volume":"75","author":"Anel","year":"2011","journal-title":"Theriogenology"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"354","DOI":"10.1016\/j.anireprosci.2009.08.005","article-title":"Ability of Catalonian donkey sperm to penetrate zona pellucida-free bovine oocytes matured in vitro","volume":"118","author":"Taberner","year":"2010","journal-title":"Anim. Reprod. Sci."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1067","DOI":"10.1016\/j.theriogenology.2014.01.033","article-title":"Evaluation of sperm subpopulation structure in relation to in vitro sperm-oocyte interaction of frozen-thawed semen from Holstein bulls","volume":"81","author":"Ferraz","year":"2014","journal-title":"Theriogenology"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"2415","DOI":"10.1095\/biolreprod.114.118539","article-title":"The Paradoxical Relationship Between Stallion Fertility and Oxidative Stress","volume":"91","author":"Gibb","year":"2014","journal-title":"Biol. Reprod."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1111\/j.1439-0531.2012.02028.x","article-title":"Seminal Plasma Proteins as Modulators of the Sperm Function and Their Application in Sperm Biotechnologies","volume":"47","author":"Caballero","year":"2012","journal-title":"Reprod. Domest. Anim."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"237","DOI":"10.1007\/s00441-020-03181-1","article-title":"Porcine model for the study of sperm capacitation, fertilization and male fertility","volume":"380","author":"Zigo","year":"2020","journal-title":"Cell Tissue Res."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"242","DOI":"10.1002\/j.1939-4640.1998.tb01994.x","article-title":"The molecular basis of sperm capacitation","volume":"19","author":"Visconti","year":"1998","journal-title":"J. Androl."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"590","DOI":"10.1095\/biolreprod61.3.590","article-title":"Bovine seminal plasma phospholipid-binding proteins stimulate phospholipid efflux from epididymal sperm","volume":"61","author":"Therien","year":"1999","journal-title":"Biol. Reprod."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"24","DOI":"10.1016\/S1388-1981(00)00080-9","article-title":"Characteristics of the cholesterol efflux induced by novel seminal phospholipid-binding proteins","volume":"1487","author":"Moreau","year":"2000","journal-title":"Biochim. Biophys. Acta Mol. Cell Biol. Lipids"},{"key":"ref_58","first-page":"332","article-title":"Isolation and characterization of phospholipid-binding proteins from goat seminal plasma","volume":"66","author":"Manjunath","year":"2002","journal-title":"Biol. Reprod."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"1447","DOI":"10.1016\/j.theriogenology.2015.07.035","article-title":"Use of combinations of in vitro quality assessments to predict fertility of bovine semen","volume":"84","author":"Sellem","year":"2015","journal-title":"Theriogenology"},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"197","DOI":"10.1071\/RDv24n1Ab171","article-title":"In vitro fertilization using frozen-thawed sexed semen treated with recombinant heparin-binding proteins","volume":"24","author":"Kjelland","year":"2012","journal-title":"Reprod. Fertil. Dev."},{"key":"ref_61","doi-asserted-by":"crossref","unstructured":"Pena, F.J., O\u2019Flaherty, C., Rodriguez, J.M.O., Cano, F.E.M., Gaitskell-Phillips, G.L., Gil, M.C., and Ferrusola, C.O. (2019). Redox Regulation and Oxidative Stress: The Particular Case of the Stallion Spermatozoa. Antioxidants, 8.","DOI":"10.3390\/antiox8110567"},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"481","DOI":"10.1016\/S0378-4320(00)00099-3","article-title":"The causes of reduced fertility with cryopreserved semen","volume":"60","author":"Watson","year":"2000","journal-title":"Anim. Reprod. Sci."},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"2407","DOI":"10.1242\/dev.127.11.2407","article-title":"The capacitating agent bicarbonate induces protein kinase A-dependent changes in phospholipid transbilayer behavior in the sperm plasma membrane","volume":"127","author":"Gadella","year":"2000","journal-title":"Development"},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"461","DOI":"10.1002\/j.1939-4640.1997.tb01953.x","article-title":"Premature capacitation of bovine spermatozoa is initiated by cryopreservation","volume":"18","author":"Cormier","year":"1997","journal-title":"J. Androl."},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1016\/j.cryobiol.2011.08.002","article-title":"Cryopreservation-induced alterations in protein tyrosine phosphorylation of spermatozoa from different portions of the boar ejaculate","volume":"63","author":"Kumaresan","year":"2011","journal-title":"Cryobiology"},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1530\/reprod\/120.1.143","article-title":"Reproductive characteristics of the African pygmy hedgehog, Atelerix albiventris","volume":"120","author":"Bedford","year":"2000","journal-title":"J. Reprod. Fertil."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1111\/j.1439-0531.2011.01818.x","article-title":"Effect of Cryopreservation on Apoptotic-like Events and its Relationship with Cryocapacitation of Buffalo (Bubalus bubalis) Sperm","volume":"47","author":"Kadirvel","year":"2012","journal-title":"Reprod. Domest. Anim."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"280","DOI":"10.1016\/j.anireprosci.2012.10.027","article-title":"Equine spermatozoa stored in the epididymis for up to 96 h at 4 degrees C can be successfully cryopreserved and maintain their fertilization capacity","volume":"136","author":"Vieira","year":"2013","journal-title":"Anim. Reprod. Sci."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"R181","DOI":"10.1530\/REP-18-0541","article-title":"Update on mammalian sperm capacitation: How much does the horse differ from other species","volume":"157","author":"Leemans","year":"2019","journal-title":"Reproduction"},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"1208","DOI":"10.1095\/biolreprod.102.011106","article-title":"Phosphorylation of protein tyrosine residues in fresh and cryopreserved stallion spermatozoa under capacitating conditions","volume":"68","author":"Pommer","year":"2003","journal-title":"Biol. Reprod."},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"424","DOI":"10.1095\/biolreprod.106.055624","article-title":"Induction of epididymal boar sperm capacitation by pB1 and BSP-A1\/-A2 proteins, members of the BSP protein family","volume":"76","author":"Lusignan","year":"2007","journal-title":"Biol. Reprod."},{"key":"ref_72","doi-asserted-by":"crossref","first-page":"1372","DOI":"10.1095\/biolreprod52.6.1372","article-title":"Phosphatidylcholine-binding proteins of bovine seminal plasma modulate capacitation of spermatozoa by heparin","volume":"52","author":"Therien","year":"1995","journal-title":"Biol. Reprod."},{"key":"ref_73","doi-asserted-by":"crossref","first-page":"84","DOI":"10.1046\/j.1365-2605.2002.00330.x","article-title":"Capacitation status and in vitro fertility of boar spermatozoa: Effects of seminal plasma, cumulus-oocyte-complexes-conditioned medium and hyaluronan","volume":"25","author":"Suzuki","year":"2002","journal-title":"Int. J. Androl."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"416","DOI":"10.2164\/jandrol.106.001826","article-title":"Effects of seminal plasma on cooling-induced capacitative changes in boar sperm","volume":"28","author":"Vadnais","year":"2007","journal-title":"J. Androl."},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"R243","DOI":"10.1530\/REP-18-0627","article-title":"Ram seminal plasma and its functional proteomic assessment","volume":"157","author":"Leahy","year":"2019","journal-title":"Reproduction"},{"key":"ref_76","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1016\/j.jprot.2013.05.029","article-title":"Proteomic characterization and cross species comparison of mammalian seminal plasma","volume":"91","author":"Druart","year":"2013","journal-title":"J. Proteom."},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"7560","DOI":"10.1073\/pnas.78.12.7560","article-title":"Timing of fertilization in mammals\u2014Sperm cholesterol phospholipid ratio as a determinant of the capacitation interval","volume":"78","author":"Davis","year":"1981","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1093\/oxfordjournals.humrep.a137045","article-title":"Variation in the cholesterol phospholipid ratio in human spermatozoa and its relationship with capacitation","volume":"5","author":"Hoshi","year":"1990","journal-title":"Hum. Reprod."},{"key":"ref_79","doi-asserted-by":"crossref","first-page":"707","DOI":"10.1093\/biolreprod\/52.monograph_series1.707","article-title":"Evaluating Capacitation of Stallion Spermatozoa Obtained from the Mare\u2019s Reproductive Tract","volume":"52","author":"Tischner","year":"1995","journal-title":"Biol. Reprod."},{"key":"ref_80","first-page":"193","article-title":"Aggregated forms of heparin-binding and non-heparin-binding proteins of boar seminal plasma and their binding properties","volume":"45","author":"Manaskova","year":"1999","journal-title":"Folia Biol."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/21\/17\/6415\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:06:18Z","timestamp":1760177178000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/21\/17\/6415"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,9,3]]},"references-count":80,"journal-issue":{"issue":"17","published-online":{"date-parts":[[2020,9]]}},"alternative-id":["ijms21176415"],"URL":"https:\/\/doi.org\/10.3390\/ijms21176415","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,9,3]]}}}