Register      Login
Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Effect of spermatozoa motility hyperactivation factors and gamete coincubation duration on in vitro bovine embryo development using flow cytometrically sorted spermatozoa

Luis B. Ferré A F , Yanina Bogliotti B , James L. Chitwood B , Cristóbal Fresno C , Hugo H. Ortega D , Michael E. Kjelland E and Pablo J. Ross B F
+ Author Affiliations
- Author Affiliations

A Instituto Nacional de Tecnología Agropecuaria (INTA), Ruta Nacional 34, Km 227, Rafaela (S2300ICB), Santa Fe, Argentina.

B Department of Animal Science, One Shields Avenue, University of California, Davis, CA 95616, USA.

C Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)/Facultad de Ingeniería, Universidad Católica de Córdoba, Av. Armada Argentina 3555, Córdoba (X5016DHK), Argentina.

D Facultad de Ciencias Veterinarias, Universidad Nacional del Litoral/CONICET, R.P. Kreder 2805, Esperanza (S3080HOF), Santa Fe, Argentina.

E Conservation, Genetics and Biotech, LLC, P.O. Box 821522, Vicksburg, MS 39182, USA.

F Corresponding authors. Emails: ferre.luis@inta.gob.ar; pross@ucdavis.edu

Reproduction, Fertility and Development 29(4) 805-814 https://doi.org/10.1071/RD15289
Submitted: 17 July 2015  Accepted: 1 December 2015   Published: 24 February 2016

Abstract

The aim of the present study was to evaluate the effects of sperm motility enhancers and different IVF times on cleavage, polyspermy, blastocyst formation, embryo quality and hatching ability. In Experiment 1, sex-sorted X chromosome-bearing Bos taurus spermatozoa were incubated for 30 min before 18 h fertilisation with hyperactivating factors, namely 10 mM caffeine (CA), 5 mM theophylline (TH), 10 mM caffeine and 5 mM theophylline (CA + TH); and untreated spermatozoa (control). In Experiment 2, matured B. taurus oocytes were fertilised using a short (8 h) or standard (18 h) fertilisation length, comparing two different fertilisation media, namely synthetic oviducal fluid (SOF) fertilisation medium (SOF-FERT) and M199 fertilisation medium (M199-FERT). Cleavage and blastocyst formation rates were significantly higher in the CA + TH group (77% and 27%, respectively) compared with the control group (71% and 21%, respectively). Cleavage rates and blastocyst formation were significantly lower for the shortest fertilisation time (8 h) in M199-FERT medium (42% and 12%, respectively). The SOF-FERT medium with an 8 h fertilisation time resulted in the highest cleavage rates and blastocyst formation (74% and 29%, respectively). The SOF-FERT medium produced the highest embryo quality (50% Grade 1) and hatching rate (66%). Motility enhancers did not affect polyspermy rates, whereas polyspermy was affected when fertilisation length was extended from 8 h (3%) to 18 h (9%) and in M199-FERT (14%) compared with SOF-FERT (6%). We conclude that adding the motility enhancers CA and TH to sex sorted spermatozoa and Tyrode’s albumin lactate pyruvate (TALP)-Sperm can improve cleavage and embryo development rates without increasing polyspermy. In addition, shortening the oocyte–sperm coincubation time (8 h) resulted in similar overall embryo performance rates compared with the prolonged (18 h) interval.

Additional keywords: IVF, sexed spermatozoa.


References

Agarwal, A., Said, T. M., Bedaiwy, M. A., Banerjee, J., and Alvarez, J. G. (2006). Oxidative stress in an assisted reproductive techniques setting. Fertil. Steril. 86, 503–512.
Oxidative stress in an assisted reproductive techniques setting.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28XhtFCqs7bJ&md5=b6fdab049392227ae5333fc33f3f2819CAS | 16860798PubMed |

Aitken, R. J., and Henkel, R. R. (2011). Sperm cell biology: current perspectives and future prospects. Asian J. Androl. 13, 3–5.
Sperm cell biology: current perspectives and future prospects.Crossref | GoogleScholarGoogle Scholar | 21102477PubMed |

Aitken, R. J., and Nixon, B. (2013). Sperm capacitation: a distant landscape glimpsed but unexplored. Mol. Hum. Reprod. 19, 785–793.
Sperm capacitation: a distant landscape glimpsed but unexplored.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXhvVOgurfM&md5=d2d6293be340a5c0d8dfeafc53dba6f0CAS | 24071444PubMed |

Aitken, R. J., Jones, K. T., and Robertson, S. A. (2012). Reactive oxygen species and sperm function: in sickness and in health. J. Androl. 33, 1096–1106.
Reactive oxygen species and sperm function: in sickness and in health.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXlvFaqsQ%3D%3D&md5=668369c26335f232861c4c306de7f315CAS | 22879525PubMed |

Amann, R. P. (1999). Issues affecting commercialization of sexed sperm. Theriogenology 52, 1441–1457.
Issues affecting commercialization of sexed sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3c7pvFalsQ%3D%3D&md5=e48a2f0f64091187ff88bae72e435e02CAS | 10735088PubMed |

Austin, C. R. (1952). The capacitation of the mammalian sperm. Nature 170, 326.
The capacitation of the mammalian sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaG3s%2FhslChsw%3D%3D&md5=406d6dcc4515c7fc16d805f2a57b0648CAS | 12993150PubMed |

Bailey, J. L. (2010). Factors regulating sperm capacitation. Syst. Biol. Reprod. Med. 56, 334–348.
Factors regulating sperm capacitation.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhtFOitLjE&md5=e6ab0abc64c8df57945eaf6aa73676caCAS | 20849222PubMed |

Baker, M. A., and Aitken, R. J. (2004). The importance of redox regulated pathways in sperm cell biology. Mol. Cell. Endocrinol. 216, 47–54.
The importance of redox regulated pathways in sperm cell biology.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXjsVens70%3D&md5=35c4ea81f6cf98b7838ea2634b82c8c6CAS | 15109744PubMed |

Bansal, A. K., and Bilaspuri, G. S. (2011). Impacts of oxidative stress and antioxidants on semen functions. Vet. Med. Int. 2011, Article ID 686137.

Barceló-Fimbres, M., Campos-Chillón, L. F., and Seidel, G. E. (2011). In vitro fertilization using non-sexed and sexed bovine sperm: sperm concentration, sorter pressure, and bull effects. Reprod. Domest. Anim. 46, 495–502.
In vitro fertilization using non-sexed and sexed bovine sperm: sperm concentration, sorter pressure, and bull effects.Crossref | GoogleScholarGoogle Scholar | 20946538PubMed |

Bedford, J. M. (1970). Sperm capacitation and fertilization in mammals. Biol. Reprod. Suppl. 2, 128–158.
Sperm capacitation and fertilization in mammals.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD38rms1ynsA%3D%3D&md5=de8ba708bab58737bbdb303e5a8a9188CAS | 12254592PubMed |

Begley, A. J., and Quinn, P. (1982). Decapacitation factors in semen. Clin. Reprod. Fertil. 1, 167–175.
| 1:STN:280:DyaL2c%2FlsV2isA%3D%3D&md5=8fa437a64da461283b64ae09d84f7f1eCAS | 7187265PubMed |

Berland, M., Frei, M., Peralta, O., and Ratto, M. (2011). Time exposure period of bovine oocytes to sperm in relation to embryo development rate and quality. ISRN Vet. Sci. 2011, Article ID 257627.
Time exposure period of bovine oocytes to sperm in relation to embryo development rate and quality.Crossref | GoogleScholarGoogle Scholar |

Biggers, J. D., McGinnis, L. K., and Raffin, M. (2000). Amino acids and preimplantation development of the mouse in protein-free potassium simplex optimized medium. Biol. Reprod. 63, 281–293.
Amino acids and preimplantation development of the mouse in protein-free potassium simplex optimized medium.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3cXktl2rsbw%3D&md5=3731fb796ae8f9769cee6fb167b5ab18CAS | 10859270PubMed |

Bird, J. M., Carey, S., and Houghton, J. A. (1989). Molitity and acrosomal changes in ionophore-treated bovine spermatozoa and their relationship with in vitro penetration of zone-free hamster oocytes. Theriogenology 32, 227–242.
Molitity and acrosomal changes in ionophore-treated bovine spermatozoa and their relationship with in vitro penetration of zone-free hamster oocytes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1MXmtFamsLY%3D&md5=9c2dc0a39491594cb89ebf88e0202601CAS | 16726670PubMed |

Brackett, B. G., and Oliphant, G. (1975). Capacitation of rabbit spermatozoa in vitro. Biol. Reprod. 12, 260–274.
Capacitation of rabbit spermatozoa in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaE28Xnt1WltQ%3D%3D&md5=8bdb4d5b8aa38c3c50168ef421c41016CAS | 1122333PubMed |

Breininger, E., Cetica, P. D., and Beconi, M. T. (2010). Capacitation inducers act through diverse intracellular mechanisms in cryopreserved bovine sperm. Theriogenology 74, 1036–1049.
Capacitation inducers act through diverse intracellular mechanisms in cryopreserved bovine sperm.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhtFWqtLnE&md5=89eea839bc3270e1514723d709415e2bCAS | 20580081PubMed |

Chatterjee, S., and Gagnon, C. (2001). Production of reactive oxygen species by spermatozoa undergoing cooling, freezing, and thawing. Mol. Reprod. Dev. 59, 451–458.
Production of reactive oxygen species by spermatozoa undergoing cooling, freezing, and thawing.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXltVWmsb4%3D&md5=d8d1459a0651eef5d9c9d78aaae63d27CAS | 11468782PubMed |

Coy, P., Garcia-Vazquez, F. A., Visconti, P. E., and Aviles, M. (2012). Roles of the oviduct in mammalian fertilization. Reproduction 144, 649–660.
Roles of the oviduct in mammalian fertilization.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XhvVOitb%2FP&md5=c9baa99fe5b5870f257a31787daaa3bfCAS | 23028122PubMed |

Cran, D. G. (2007). XY sperm separation and use in artificial insemination and other ARTs. Soc. Reprod. Fertil. Suppl. 65, 475–491.
| 17644986PubMed |

Dalvit, G. C., Cetica, P. D., Pintos, L. N., and Beconi, M. T. (2005). Reactive oxygen species in bovine embryo in vitro production. Biocell 29, 209–212.
| 1:CAS:528:DC%2BD2MXhtFSns73M&md5=654b4b23be6c3704bda4163fae9694feCAS | 16187501PubMed |

De Vries, A., Overton, M., Fetrow, J., Leslie, K., Eicker, S., and Rogers, G. (2008). Exploring the impact of sexed semen on the structure of the dairy industry. J. Dairy Sci. 91, 847–856.
Exploring the impact of sexed semen on the structure of the dairy industry.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXhsVaitLw%3D&md5=cacca6ea9f61f4bf0d0d14e560ca37deCAS | 18218773PubMed |

Di Rienzo, J., Casanoves, F., Balzarini, M., Gonzalez, L., Tablada, M., and Robledo, C. (2011). InfoStat v2011. (InfoStat Group, College of Agricultural Sciences, National University of Córdoba: Argentina.)

Enkhmaa, D., Kasai, T., and Hoshi, K. (2009). Long-time exposure of mouse embryos to the sperm produces high levels of reactive oxygen species in culture medium and relates to poor embryo development. Reprod. Domest. Anim. 44, 634–637.
Long-time exposure of mouse embryos to the sperm produces high levels of reactive oxygen species in culture medium and relates to poor embryo development.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXhtVaiurnI&md5=fb43067781afbc8297ecde66659f62ebCAS | 19019063PubMed |

Evans, G., Hollinshead, F. K., and Maxwell, W. M. (2004). Preservation and artificial insemination of sexed semen in sheep. Reprod. Fertil. Dev. 16, 455–464.
Preservation and artificial insemination of sexed semen in sheep.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2cvht1WnsA%3D%3D&md5=a62b111dcb07e4173aa4fd7b050980b9CAS | 15315744PubMed |

Ferré, L. B., Bogliotti, Y., Chitwood, J. L., Fresno, C., Ortega, H. H., Kjelland, M. E., and Ross, P. J. (2015). Comparison of different fertilisation media for an in vitro maturation–fertilisation–culture system using flow-cytometrically sorted X chromosome-bearing spermatozoa for bovine embryo production. Reprod. Fertil. Dev. , .
Comparison of different fertilisation media for an in vitro maturation–fertilisation–culture system using flow-cytometrically sorted X chromosome-bearing spermatozoa for bovine embryo production.Crossref | GoogleScholarGoogle Scholar | 25966894PubMed |

Florman, H. M., and Ducibella, T. (2006). Mammalian fertilization. In ‘The Physiology of Reproduction’. 3rd edn. (Eds E. Knobil and J. D. Neill.) pp. 55–112. (Elsevier Academic Press Publications: San Diego, CA)

Fraser, L. R. (1998). Sperm capacitation and the acrosome reaction. Hum. Reprod. 13, 9–19.
Sperm capacitation and the acrosome reaction.Crossref | GoogleScholarGoogle Scholar | 9663766PubMed |

Fraser, L. R. (2010). The ‘switching on’ of mammalian spermatozoa: molecular events involved in promotion and regulation of capacitation. Mol. Reprod. Dev. 77, 197–208.
| 1:CAS:528:DC%2BC3cXotFSnuw%3D%3D&md5=a9633fb11ad5be06fe4d73cdf05d725aCAS | 19908247PubMed |

Garner, D. L. (2001). Sex-sorting mammalian sperm: concept to application in animals. J. Androl. 22, 519–526.
| 1:STN:280:DC%2BD38%2FitFarsg%3D%3D&md5=dd71ae48cd468eaeedf23ec77ce9e342CAS | 11451346PubMed |

Garner, D. L. (2006). Flow cytometric sexing of mammalian sperm. Theriogenology 65, 943–957.
Flow cytometric sexing of mammalian sperm.Crossref | GoogleScholarGoogle Scholar | 16242764PubMed |

Garner, D. L. (2009). Hoechst 33342: the dye that enabled differentiation of living X- and Y-chromosome bearing mammalian sperm. Theriogenology 71, 11–21.
Hoechst 33342: the dye that enabled differentiation of living X- and Y-chromosome bearing mammalian sperm.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXhsVCksr%2FL&md5=63996b58e460aefd55ae4e76f029946aCAS | 18952273PubMed |

Garner, D. L., Evans, K. M., and Seidel, G. E. (2013). Sex-sorting sperm using flow cytometry/cell sorting. Methods Mol. Biol. 927, 279–295.
Sex-sorting sperm using flow cytometry/cell sorting.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXit1SmsrY%3D&md5=cc695225fe27c4ca12ced0d285bc2dd8CAS | 22992923PubMed |

Gordon, I. R. (2003). Capacitating bovine sperm. In ‘Laboratory Production of Cattle Embryos’. 2nd edn. (Ed. I. R. Gordon.) pp. 158–175. (CABI Pub.: Cambridge, MA)

Gosálvez, J., Ramirez, M. A., López-Fernández, C., Crespo, F., Evans, K. M., Kjelland, M. E., and Moreno, J. F. (2011a). Sex-sorted bovine spermatozoa and DNA damage: I. Static features. Theriogenology 75, 197–205.
Sex-sorted bovine spermatozoa and DNA damage: I. Static features.Crossref | GoogleScholarGoogle Scholar | 20932559PubMed |

Gosálvez, J., Ramirez, M. A., López-Fernández, C., Crespo, F., Evans, K. M., Kjelland, M. E., and Moreno, J. F. (2011b). Sex-sorted bovine spermatozoa and DNA damage: II. Dynamic features. Theriogenology 75, 206–211.
Sex-sorted bovine spermatozoa and DNA damage: II. Dynamic features.Crossref | GoogleScholarGoogle Scholar | 21040960PubMed |

Guérin, P., El Mouatassim, S., and Ménézo, Y. (2001). Oxidative stress and protection against reactive oxygen species in the pre-implantation embryo and its surroundings. Hum. Reprod. Update 7, 175–189.
Oxidative stress and protection against reactive oxygen species in the pre-implantation embryo and its surroundings.Crossref | GoogleScholarGoogle Scholar | 11284661PubMed |

Henkel, R. (2012). Sperm preparation: state-of-the-art–physiological aspects and application of advanced sperm preparation methods. Asian J. Androl. 14, 260–269.
Sperm preparation: state-of-the-art–physiological aspects and application of advanced sperm preparation methods.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XjtlGqtb4%3D&md5=d8ddea772006fddb993fd17548d16e11CAS | 22138904PubMed |

Henkel, R. R., and Schill, W. B. (2003). Sperm preparation for ART. Reprod. Biol. Endocrinol. 1, 108.
Sperm preparation for ART.Crossref | GoogleScholarGoogle Scholar | 14617368PubMed |

Ho, H.-C., and Suarez, S. S. (2001a). An inositol 1,4,5-trisphosphate receptor-gated intracellular Ca2+ store is involved in regulating sperm hyperactivated motility. Biol. Reprod. 65, 1606–1615.
An inositol 1,4,5-trisphosphate receptor-gated intracellular Ca2+ store is involved in regulating sperm hyperactivated motility.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXnvVersb0%3D&md5=8766b0703c2c071d3f65bead454213b0CAS | 11673282PubMed |

Ho, H. C., and Suarez, S. S. (2001b). Hyperactivation of mammalian spermatozoa: function and regulation. Reproduction 122, 519–526.
Hyperactivation of mammalian spermatozoa: function and regulation.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXotFGltb8%3D&md5=27198eb881c4ab0908d838dec5e1e761CAS | 11570958PubMed |

Hohenboken, W. D. (1999). Applications of sexed semen in cattle production. Theriogenology 52, 1421–1433.
Applications of sexed semen in cattle production.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3c7pvFalsw%3D%3D&md5=644bf9d2bfc616b2ef1d08121c969099CAS | 10735086PubMed |

Hollinshead, F. K., Gillan, L., O’Brien, J. K., Evans, G., and Maxwell, W. M. C. (2003). In vitro and in vivo assessment of functional capacity of flow cytometrically sorted ram spermatozoa after freezing and thawing. Reprod. Fertil. Dev. 15, 351–359.
In vitro and in vivo assessment of functional capacity of flow cytometrically sorted ram spermatozoa after freezing and thawing.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2c%2Fos1OksQ%3D%3D&md5=6e36f3058b5755a250c4e4de8d50fc3bCAS | 14975233PubMed |

Johnson, L. A. (1991). Sex preselection in swine: altered sex ratios in offspring following surgical insemination of flow sorted X- and Y-bearing sperm. Reprod. Domest. Anim. 26, 309–314.
Sex preselection in swine: altered sex ratios in offspring following surgical insemination of flow sorted X- and Y-bearing sperm.Crossref | GoogleScholarGoogle Scholar |

Kang, S. S., Koyama, K., Huang, W., Yanagawa, Y., Takahashi, Y., and Nagano, M. (2015). Addition of d-penicillamine, hypotaurine, and epinephrine (PHE) mixture to IVF medium maintains motility and longevity of bovine sperm and enhances stable production of blastocysts in vitro. J. Reprod. Dev. 61, 99–105.
Addition of d-penicillamine, hypotaurine, and epinephrine (PHE) mixture to IVF medium maintains motility and longevity of bovine sperm and enhances stable production of blastocysts in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC2MXosV2lt7k%3D&md5=e87d9d05d7ba3ea652436057bee3c862CAS | 25501343PubMed |

Kattera, S., and Chen, C. (2003). Short coincubation of gametes in in vitro fertilization improves implantation and pregnancy rates: a prospective, randomized, controlled study. Fertil. Steril. 80, 1017–1021.
Short coincubation of gametes in in vitro fertilization improves implantation and pregnancy rates: a prospective, randomized, controlled study.Crossref | GoogleScholarGoogle Scholar | 14556826PubMed |

Kay, V. J., and Robertson, L. (1998). Hyperactivated motility of human spermatozoa: a review of physiological function and application in assisted reproduction. Hum. Reprod. Update 4, 776–786.
Hyperactivated motility of human spermatozoa: a review of physiological function and application in assisted reproduction.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK1M7ps1Wjtw%3D%3D&md5=927258a4af4ff682465c4ca31ff9951aCAS | 10098469PubMed |

Kochhar, H. S., Kochhar, K. P., Basrur, P. K., and King, W. A. (2003). Influence of the duration of gamete interaction on cleavage, growth rate and sex distribution of in vitro produced bovine embryos. Anim. Reprod. Sci. 77, 33–49.
Influence of the duration of gamete interaction on cleavage, growth rate and sex distribution of in vitro produced bovine embryos.Crossref | GoogleScholarGoogle Scholar | 12654526PubMed |

Koyama, K., Kang, S. S., Huang, W., Yanagawa, Y., Takahashi, Y., and Nagano, M. (2014). Estimation of the optimal timing of fertilization for embryo development of in vitro-matured bovine oocytes based on the times of nuclear maturation and sperm penetration. J. Vet. Med. Sci. 76, 653–659.
Estimation of the optimal timing of fertilization for embryo development of in vitro-matured bovine oocytes based on the times of nuclear maturation and sperm penetration.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC2cXhtFCrtrfJ&md5=bfab96e706e3574e72449bfaab3b7677CAS | 24430663PubMed |

Lanzafame, F., Chapman, M. G., Guglielmino, A., Gearon, C. M., and Forman, R. G. (1994). Pharmacological stimulation of sperm motility. Hum. Reprod. 9, 192–199.
| 1:CAS:528:DyaK2cXkslarsL8%3D&md5=f463fa821b6ea4864572dec47ca21a6aCAS | 8027272PubMed |

Lin, S. P., Lee, R. K., Su, J. T., Lin, M. H., and Hwu, Y. M. (2000). The effects of brief gamete co-incubation in human in vitro fertilization. J. Assist. Reprod. Genet. 17, 344–348.
The effects of brief gamete co-incubation in human in vitro fertilization.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3cvptFGqtg%3D%3D&md5=f749de317cf0eb0ea1c3f103370aa8bfCAS | 11042832PubMed |

Lindsey, A. C., Bruemmer, J. E., and Squires, E. L. (2001). Low dose insemination of mares using non-sorted and sex-sorted sperm. Anim. Reprod. Sci. 68, 279–289.
Low dose insemination of mares using non-sorted and sex-sorted sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3MnpslKnuw%3D%3D&md5=8ded3e629eb117f385055a4a520042d8CAS | 11744272PubMed |

Long, C. R., Damiani, P., Pinto-Correia, C., MacLean, R. A., Duby, R. T., and Robl, J. M. (1994). Morphology and subsequent development in culture of bovine oocytes matured in vitro under various conditions of fertilization. J. Reprod. Fertil. 102, 361–369.
Morphology and subsequent development in culture of bovine oocytes matured in vitro under various conditions of fertilization.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2MXjtFyns7w%3D&md5=c721ab666d5cb1864745438bbc45101bCAS | 7861389PubMed |

Lopes, A. S., Lane, M., and Thompson, J. G. (2010). Oxygen consumption and ROS production are increased at the time of fertilization and cell cleavage in bovine zygotes. Hum. Reprod. 25, 2762–2773.
Oxygen consumption and ROS production are increased at the time of fertilization and cell cleavage in bovine zygotes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhtlShtb7O&md5=d95363a45e51c9cf84b25b6396a619f2CAS | 20823113PubMed |

Loughlin, K. R., and Agarwal, A. (1992). Use of theophylline to enhance sperm function. Arch. Androl. 28, 99–103.
Use of theophylline to enhance sperm function.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK38XktVSrt7w%3D&md5=3a6f40c5fc3d68c654b879ce3e093504CAS | 1520041PubMed |

Lu, K. H., and Seidel, G. E. (2004). Effects of heparin and sperm concentration on cleavage and blastocyst development rates of bovine oocytes inseminated with flow cytometrically-sorted sperm. Theriogenology 62, 819–830.
Effects of heparin and sperm concentration on cleavage and blastocyst development rates of bovine oocytes inseminated with flow cytometrically-sorted sperm.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXls1aisbo%3D&md5=a89eab921e7a4076a2909bec11c4486fCAS | 15251233PubMed |

Lu, K. H., Cran, D. G., and Seidel, G. E. (1999). In vitro fertilization with flow-cytometrically-sorted bovine sperm. Theriogenology 52, 1393–1405.
In vitro fertilization with flow-cytometrically-sorted bovine sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3c7pvFamuw%3D%3D&md5=487d491bb1856c86d7db84060365d610CAS | 10735084PubMed |

Marquez, B., and Suarez, S. S. (2007). Bovine sperm hyperactivation is promoted by alkaline-stimulated Ca2+ influx. Biol. Reprod. 76, 660–665.
Bovine sperm hyperactivation is promoted by alkaline-stimulated Ca2+ influx.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXjsFCmsr8%3D&md5=57deacf8bf12ea3a704eaf231f3dab27CAS | 17182893PubMed |

Mocé, E., Graham, J. K., and Schenk, J. L. (2006). Effect of sex-sorting on the ability of fresh and cryopreserved bull sperm to undergo an acrosome reaction. Theriogenology 66, 929–936.
Effect of sex-sorting on the ability of fresh and cryopreserved bull sperm to undergo an acrosome reaction.Crossref | GoogleScholarGoogle Scholar | 16564078PubMed |

Momozawa, K., and Fukuda, Y. (2003). Caffeine in fertilization medium is not essential for bovine IVF by fully capacitated spermatozoa. J. Reprod. Dev. 49, 507–512.
Caffeine in fertilization medium is not essential for bovine IVF by fully capacitated spermatozoa.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXhvFahsrw%3D&md5=db2ff0df7d60383f63c0dcec46a2af74CAS | 14967902PubMed |

Mortimer, D., Barratt, C. L. R., Björndahl, L., de Jager, C., Jequier, A. M., and Muller, C. H. (2013). What should it take to describe a substance or product as ‘sperm-safe’. Hum. Reprod. Update 19, i1–i45.
What should it take to describe a substance or product as ‘sperm-safe’.Crossref | GoogleScholarGoogle Scholar | 23552271PubMed |

Nedambale, T. L., Du, F., Xu, J., Chaubal, S. A., Dinnyes, A., Groen, W., Faber, D., Dobrinsky, J. R., Yang, X., and Tian, X. C. (2006). Prolonging bovine sperm–oocyte incubation in modified medium 199 improves embryo development rate and the viability of vitrified blastocysts. Theriogenology 66, 1951–1960.
Prolonging bovine sperm–oocyte incubation in modified medium 199 improves embryo development rate and the viability of vitrified blastocysts.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD28nlsF2isA%3D%3D&md5=ac7f1c8c6fb99b9d0cb9adff65cb61adCAS | 16787658PubMed |

Niwa, K., and Ohgoda, O. (1988). Synergistic effect of caffeine and heparin on in-vitro fertilization of cattle oocytes matured in culture. Theriogenology 30, 733–741.
Synergistic effect of caffeine and heparin on in-vitro fertilization of cattle oocytes matured in culture.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1MXltFCisQ%3D%3D&md5=97c5f0745afd99a2b7fd604b3d0ee72aCAS | 16726515PubMed |

Numabe, T., Oikawa, T., Kikuchi, T., and Horuchi, T. (2001). Pentoxifylline improves in vitro fertilization and subsequent development of bovine oocytes. Theriogenology 56, 225–233.
Pentoxifylline improves in vitro fertilization and subsequent development of bovine oocytes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXlvVyntLk%3D&md5=e4c0c00a558a89e2b943ba4d234b2c81CAS | 11480615PubMed |

Park, C. K., Ohgoda, O., and Niwa, K. (1989). Penetration of bovine follicular oocytes by frozen–thawed spermatozoa in the presence of caffeine and heparin. J. Reprod. Fertil. 86, 577–582.
Penetration of bovine follicular oocytes by frozen–thawed spermatozoa in the presence of caffeine and heparin.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1MXkvFCls7Y%3D&md5=09a103f634f4effb9bd67db8bc2b7b85CAS | 2760886PubMed |

Parrish, J. J. (2014). Bovine in vitro fertilization: in vitro oocyte maturation and sperm capacitation with heparin. Theriogenology 81, 67–73.
Bovine in vitro fertilization: in vitro oocyte maturation and sperm capacitation with heparin.Crossref | GoogleScholarGoogle Scholar | 24274411PubMed |

Parrish, J. J., Susko-Parrish, J. L., Leibfried-Rutledge, M. L., Critser, E. S., Eyestone, W. H., and First, N. L. (1986). Bovine in vitro fertilization with frozen–thawed semen. Theriogenology 25, 591–600.
Bovine in vitro fertilization with frozen–thawed semen.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD283pvV2itQ%3D%3D&md5=9fddc262c2bd76e8223bdbbd4bc7e9a6CAS | 16726150PubMed |

Parrish, J. J., Susko-Parrish, J., Winer, M. A., and First, N. L. (1988). Capacitation of bovine sperm by heparin. Biol. Reprod. 38, 1171–1180.
Capacitation of bovine sperm by heparin.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL1cXkslWit7g%3D&md5=5f6eee533fe3840b8dccbb5ba5b04cabCAS | 3408784PubMed |

Parrish, J. J., Susko-Parrish, J. L., and First, N. L. (1989). Capacitation of bovine sperm by heparin: inhibitory effect of glucose and role of intracellular pH. Biol. Reprod. 41, 683–699.
Capacitation of bovine sperm by heparin: inhibitory effect of glucose and role of intracellular pH.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3cXmvFOmsQ%3D%3D&md5=a9baa6b32aca25baafed909cdbb9289eCAS | 2620077PubMed |

R Development Core Team (2014). ‘R: A Language and Environment for Statistical Computing.’ (R Foundation for Statistical Computing: Vienna.)

Rath, D., Moench-Tegeder, G., Taylor, U., and Johnson, L. A. (2009). Improved quality of sex-sorted sperm: a prerequisite for wider commercial application. Theriogenology 71, 22–29.
Improved quality of sex-sorted sperm: a prerequisite for wider commercial application.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD1cjnt1CksQ%3D%3D&md5=b052f12eb45f7986cfc35f1166a9e083CAS | 18995893PubMed |

Rehman, N., Collins, A. R., Suh, T. K., and Wright, R. W. (1994). Effect of sperm exposure time on in vitro fertilization and embryo development of bovine oocytes matured in vitro. Theriogenology 41, 1447–1452.
Effect of sperm exposure time on in vitro fertilization and embryo development of bovine oocytes matured in vitro.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD28zgtVamtA%3D%3D&md5=b5897920940cbb66bbb6298f509b51e2CAS | 16727498PubMed |

Schenk, J. L., and Seidel, G. E. (2007). Pregnancy rates in cattle with cryopreserved sexed spermatozoa: effects of laser intensity, staining conditions and catalase. Soc. Reprod. Fertil. Suppl. 64, 165–177.
| 1:STN:280:DC%2BD2s3otlKjuw%3D%3D&md5=086add7a52cfd89b99feaf4f94df7737CAS | 17491146PubMed |

Schenk, J. L., Suh, T. K., Cran, D. G., and Seidel, G. E. (1999). Cryopreservation of flow-sorted bovine spermatozoa. Theriogenology 52, 1375–1391.
Cryopreservation of flow-sorted bovine spermatozoa.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3c7pvFamug%3D%3D&md5=969153348ef6786c32fc8c7dc25a3d99CAS | 10735083PubMed |

Seidel, G. E., and Garner, D. L. (2002). Current status of sexing mammalian spermatozoa. Reproduction 124, 733–743.
Current status of sexing mammalian spermatozoa.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3sXntVWitg%3D%3D&md5=fd55391943c27ed75ecc2e13158c6944CAS | 12537000PubMed |

Stringfellow, D. A., Givens, M. D., and Society, I. E. T. (2010) ‘Manual of the International Embryo Transfer Society: A Procedural Guide and General Information for the Use of Embryo Transfer Technology Emphasizing Sanitary Procedures.’ (International Embryo Transfer Society: Savoy, IL.)

Suarez, S. S. (2008). Control of hyperactivation in sperm. Hum. Reprod. Update 14, 647–657.
Control of hyperactivation in sperm.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXht1Ois7%2FK&md5=04cfc95d07cda10ca1f29261b1cf242bCAS | 18653675PubMed |

Suarez, S. S., and Ho, H. C. (2003). Hyperactivated motility in sperm. Reprod. Domest. Anim. 38, 119–124.
Hyperactivated motility in sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3s7nvFyitQ%3D%3D&md5=c41bb29194e949c4bac0e5390f2bdd78CAS | 12654022PubMed |

Suarez, S. S., Katz, D. F., Owen, D. H., Andrew, J. B., and Powell, R. L. (1991). Evidence for the function of hyperactivated motility in sperm. Biol. Reprod. 44, 375–381.
Evidence for the function of hyperactivated motility in sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK3M7otlSquw%3D%3D&md5=175083773da230ee0d3c5290e1adec11CAS | 2009336PubMed |

Suarez, S. S., Varosi, S. M., and Dai, X. (1993). Intracellular calcium increases with hyperactivation in intact, moving hamster sperm and oscillates with the flagellar beat cycle. Proc. Natl Acad. Sci. USA 90, 4660–4664.
Intracellular calcium increases with hyperactivation in intact, moving hamster sperm and oscillates with the flagellar beat cycle.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3sXksVKiur4%3D&md5=9c2d4a86c9d9d18a999fb95ebca80766CAS | 8506314PubMed |

Suh, T., and Schenk, J. (2003). Pressure during flow sorting of bull sperm affects post-thaw motility characteristics. Theriogenology 59, 516.

Suh, T. K., Schenk, J. L., and Seidel, G. E. (2005). High pressure flow cytometric sorting damages sperm. Theriogenology 64, 1035–1048.
High pressure flow cytometric sorting damages sperm.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2MvlvF2mtA%3D%3D&md5=53921411c541dd53a729dda03c9d055dCAS | 16125550PubMed |

Takahashi, Y., and First, N. L. (1993). In vitro fertilization of bovine oocytes in the presence of theophylline. Anim. Reprod. Sci. 34, 1–18.
In vitro fertilization of bovine oocytes in the presence of theophylline.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2cXitFersbY%3D&md5=6a46ffeaf970645221035ea222b20300CAS |

Tervit, H. R., Whittingham, D. G., and Rowson, L. E. (1972). Successful culture in vitro of sheep and cattle ova. J. Reprod. Fertil. 30, 493–497.
Successful culture in vitro of sheep and cattle ova.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaE3s%2FgvFamug%3D%3D&md5=05fb25f8136cf441849f8aef827110beCAS | 4672493PubMed |

Tsunoda, S., Kimura, N., and Fujii, J. (2014). Oxidative stress and redox regulation of gametogenesis, fertilization, and embryonic development. Reprod. Med. Biol. 13, 71–79.
Oxidative stress and redox regulation of gametogenesis, fertilization, and embryonic development.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC2cXls12qsL0%3D&md5=7f51263cbdf7c6e1fc94a538c5fa7996CAS |

Visconti, P. E., and Kopf, G. S. (1998). Regulation of protein phosphorylation during sperm capacitation. Biol. Reprod. 59, 1–6.
Regulation of protein phosphorylation during sperm capacitation.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1cXktFaksLw%3D&md5=ab59719030dd46b980e9c1282948dfe0CAS | 9674985PubMed |

Visconti, P. E., Galantino-Homer, H., Moore, G. D., Bailey, J. L., Ning, X., Fornes, M., and Kopf, G. S. (1998). The molecular basis of sperm capacitation. J. Androl. 19, 242–248.
| 1:STN:280:DyaK1c3jtVKktQ%3D%3D&md5=5af7113122eca8532ff5a4d9227643d8CAS | 9570749PubMed |

Ward, F., Enright, B., Rizos, D., Boland, M., and Lonergan, P. (2002). Optimization of in vitro bovine embryo production: effect of duration of maturation, length of gamete co-incubation, sperm concentration and sire. Theriogenology 57, 2105–2117.
Optimization of in vitro bovine embryo production: effect of duration of maturation, length of gamete co-incubation, sperm concentration and sire.Crossref | GoogleScholarGoogle Scholar | 12066869PubMed |

Weigel, K. A. (2004). Exploring the role of sexed semen in dairy production systems. J. Dairy Sci. 87, E120–E130.
Exploring the role of sexed semen in dairy production systems.Crossref | GoogleScholarGoogle Scholar |

Yanagimachi, R. (1989). Sperm capacitation and gamete interaction. J. Reprod. Fertil. Suppl. 38, 27–33.
| 1:STN:280:DyaK3c%2Fht1Shtg%3D%3D&md5=c683785960ae6dc5b686b7c27134a9e7CAS | 2677347PubMed |

Yanagimachi, R. (1994a). Fertility of mammalian spermatozoa: its development and relativity. Zygote 2, 371–372.
Fertility of mammalian spermatozoa: its development and relativity.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK283msVymtg%3D%3D&md5=339063ca10704126422b592046e241d2CAS | 8665172PubMed |

Yanagimachi, R. (1994b) Mammalian fertilization. In ‘The Physiology of Reproduction’. (Eds E. Knobil and J. D. Neill.) pp. 189–317. (Raven Press: New York.)

Yanagimachi, R. (2011). Mammalian sperm acrosome reaction: where does it begin before fertilization? Biol. Reprod. 85, 4–5.
Mammalian sperm acrosome reaction: where does it begin before fertilization?Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXotFOlsrw%3D&md5=61564d5cdccc73c330fa066bfc30d04eCAS | 21490244PubMed |

Zhang, M., Lu, K. H., and Seidel, G. E. (2003). Development of bovine embryos after in vitro fertilization of oocytes with flow cytometrically sorted, stained and unsorted sperm from different bulls. Theriogenology 60, 1657–1663.
Development of bovine embryos after in vitro fertilization of oocytes with flow cytometrically sorted, stained and unsorted sperm from different bulls.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3sXot12jsL8%3D&md5=cbe5319d437681724cf1ac91c725ae4eCAS | 14580648PubMed |