Molecular Biology Reports

, Volume 41, Issue 1, pp 217–223 | Cite as

Tentative identification of sex-specific antibodies and their application for screening bovine sperm proteins for sex-specificity

  • Wu-Cai Yang
  • Lei Sang
  • Yao Xiao
  • Hua-Lin Zhang
  • Ke-Qiong Tang
  • Li-Guo Yang
Article

Abstract

Our previous studies indicated that a purified rabbit antiserum against X-sperm contained sex-specific antibodies (SSAbs) which preferentially bound to sex-sorted X-sperm. The specificity of sex-specific antiserum was initially demonstrated using flow cytometry only, which resulted in uncertainty. In this study, the putative SSAbs against bovine X-sperm (XSSAb) were produced by a series of immunological approaches, and the effectiveness of separation of sperm using putative XSSAb was validated. Subsequently, the XSSAb was used to immunoprecipitate sex-specific proteins (SSPs) in bovine sperm, followed by two-dimensional gel electrophoresis. The results showed 7.6, 15.2 and 52.1 % of sex-sorted Y-sperm, sex-sorted X-sperm and unsorted sperm were recognized by the neutralized rabbit antisera against X-sperm, respectively. Also the purity of separation of sperm using putative XSSAb reached 74.3 % when the immunologically separated sperm were injected into oocytes. In addition, three candidate SSP sports about 30 kDa were captured by the XSSAb. Our results confirmed that the putative XSSAb contained SSAbs, and implied that these three protein sports might be SSPs in bovine X-sperm. This provides a potentially more efficient method for sorting sperm and lays a foundation for future search for SSPs.

Keywords

Sex-specific antibodies Sex-specific proteins 2-DE Sexing 

References

  1. 1.
    Johnson LA (2000) Sexing mammalian sperm for production of offspring: the state-of-the-art. Anim Reprod Sci 61:93–107CrossRefGoogle Scholar
  2. 2.
    Rath D, Johnson LA (2008) Application and commercialization of flow cytometrically sex-sorted semen. Reprod Domest Anim 2:338–346CrossRefGoogle Scholar
  3. 3.
    Ellis PJ, Yu Y, Zhang S (2011) Transcriptional dynamics of the sex chromosomes and the search for offspring sex-specific antigens in sperm. Reproduction 142(5):609–619PubMedCrossRefGoogle Scholar
  4. 4.
    Han TL, Flaherty SP, Ford JH, Matthews CD (1993) Detection of X- and Y-bearing human spermatozoa after motile sperm isolation by swim-up. Fertil Steril 60:1046–1051PubMedGoogle Scholar
  5. 5.
    Steeno O, Adimoelja A, Steeno J (1975) Separation of X- and Y-bearing human spermatozoa with the sephadex gel-filtration method. Andrologia 7:95–97PubMedCrossRefGoogle Scholar
  6. 6.
    Bennett D, Boyse EA (1973) Sex ratio in progeny of mice inseminated with sperm treated with H–Y antiserum. Nature 246:308–309PubMedCrossRefGoogle Scholar
  7. 7.
    Hendriksen PJ (1999) Do X and Y spermatozoa differ in proteins? Theriogenology 52:1295–1307PubMedCrossRefGoogle Scholar
  8. 8.
    Koundouros S, Verma P (2012) Significant enrichment of Y-bearing chromosome human spermatozoa using a modified centrifugation technique. Int J Androl 35(6):880–886PubMedCrossRefGoogle Scholar
  9. 9.
    Garner DL, Seidel GE Jr (2008) History of commercializing sexed semen for cattle. Theriogenology 69:886–895PubMedCrossRefGoogle Scholar
  10. 10.
    O’Brien JK, Steinman KJ, Robeck TR (2009) Application of sperm sorting and associated reproductive technology for wildlife management and conservation. Theriogenology 71:98–107PubMedCrossRefGoogle Scholar
  11. 11.
    Rath D, Moench-Tegeder G, Taylor U, Johnson LA (2009) Improved quality of sex-sorted sperm: a prerequisite for wider commercial application. Theriogenology 71:22–29PubMedCrossRefGoogle Scholar
  12. 12.
    Seidel GE Jr (2003) Sexing mammalian sperm-intertwining of commerce, technology, and biology. Anim Reprod Sci 79:145–156PubMedCrossRefGoogle Scholar
  13. 13.
    Blecher SR, Howie R, Li S, Detmar J, Blahut LM (1999) A new approach to immunological sexing of sperm. Theriogenology 52:1309–1321PubMedCrossRefGoogle Scholar
  14. 14.
    Hendriksen PJ, Welch GR, Grootegoed JA, Van der Lende T, Johnson LA (1996) Comparison of detergent-solubilized membrane and soluble proteins from flow cytometrically sorted X- and Y-chromosome bearing porcine spermatozoa by high resolution 2-D electrophoresis. Mol Reprod Dev 45:342–350PubMedCrossRefGoogle Scholar
  15. 15.
    Hendriksen PJ, Hoogerbrugge JW, Themmen AP, Koken MH, Hoeijmakers JH, Oostra BA, van der Lende T, Grootegoed JA (1995) Postmeiotic transcription of X and Y chromosomal genes during spermatogenesis in the mouse. Dev Biol 170:730–733PubMedCrossRefGoogle Scholar
  16. 16.
    Hamady M, Cheung THT, Resing K, Cios KJ, Knight R (2005) Key challenges in proteomics and proteoinformatics. Eng Med Biol Mag 24:34–40Google Scholar
  17. 17.
    Martin-DeLeon PA, Zhang H, Morales CR, Zhao Y, Rulon M, Barnoski BL, Chen H, Galileo DS (2005) Spam1-associated transmission ratio distortion in mice: elucidating the mechanism. Reprod Biol Endocrinol 3:32PubMedCentralPubMedCrossRefGoogle Scholar
  18. 18.
    Westbrook VA, Schoppee PD, Vanage GR, Klotz KL, Diekman AB, Flickinger CJ, Coppola MA, Herr JC (2006) Hominoid-specific SPANXA/D genes demonstrate differential expression in individuals and protein localization to a distinct nuclear envelope domain during spermatid morphogenesis. Mol Hum Reprod 12:703–716PubMedCrossRefGoogle Scholar
  19. 19.
    Mueller JL, Mahadevaiah SK, Park PJ, Warburton PE, Page DC, Turner JMA (2008) The mouse X chromosome is enriched for multicopy testis genes showing postmeiotic expression. Nat Genet 40:794–799PubMedCentralPubMedCrossRefGoogle Scholar
  20. 20.
    Véron N, Bauer H, Weisse AY, Lüder G, Werber M, Herrmann BG (2009) Retention of gene products in syncytial spermatids promotes non-Mendelian inheritance as revealed by the t complex responder. Genes Dev 23:2705–2710PubMedCrossRefGoogle Scholar
  21. 21.
    Beerli RR, Bauer MR, Buser B, Gwerder M, Muntwiler S, Maurer P, Saudan P, Bachmann MF (2008) Isolation of human monoclonal antibodies by mammalian cell display. Proc Natl Acad Sci 105:14336–14341PubMedCrossRefGoogle Scholar
  22. 22.
    Sivapurapu N, Upadhyay A, Hasegawa A, Koyama K, Gupta SK (2003) Efficacy of antibodies against Escherichia coli expressed chimeric recombinant protein encompassing multiple epitopes of zona pellucida glycoproteins to inhibit in vitro human sperm-egg binding. Mol Reprod Dev 65:309–317PubMedCrossRefGoogle Scholar
  23. 23.
    Mollova M, Djarkova T, Ivanova M, Stamenova M, Kyurkchiev S (1999) Isolation and biological characterization of boar sperm capacitation-related antigen. Am J Reprod Immunol 42:254–262PubMedCrossRefGoogle Scholar
  24. 24.
    Khan SA, Suryawanshi AR, Ranpura SA, Jadhav SV, Khole VV (2009) Identification of novel immunodominant epididymal sperm proteins using combinatorial approach. Reproduction 138:81–93PubMedCrossRefGoogle Scholar
  25. 25.
    Sang L, Yang WC, Han L, Liang AX, Hua GH, Xiong JJ, Huo LJ, Yang LG (2011) An immunological method to screen sex-specific proteins of bovine sperm. J Dairy Sci 94:2060–2070PubMedCrossRefGoogle Scholar
  26. 26.
    Pasquali C, Fialka I, Huber LA (1997) Preparative two-dimensional gel electrophoresis of membrance proteins. Electrophoresis 18:2573–2581PubMedCrossRefGoogle Scholar
  27. 27.
    Keskintepe L, Pacholczyk G, Machnicka A, Norris K, Curuk MA, Khan I, Bracket BG (2012) Bovine blastocyst development from oocytes injected with freeze-dried spermatozoa. Biol Reprod 67:409–415CrossRefGoogle Scholar
  28. 28.
    Xiao Y, Zhang HL, Ahmad S, Bai LY, Wang XM, Huo LJ, Zhang X, Li WG, Li X, Yang LG (2013) Sperm capacitation combined with removal of the sperm acrosome and plasma membrane enhances paternal nucleus remodelling and early development of bovine androgenetic embryos. Reprod Fertil Dev 25(4):624–638. doi:10.1071/RD12075 PubMedCrossRefGoogle Scholar
  29. 29.
    Li G, Chen J, Xie P, Jiang Y, Wu L, Zhang X (2011) Protein expression profiling in the zebrafish (Danio rerio) embryos exposed to the microcystin-LR. Proteomics 11:2003–2018PubMedCrossRefGoogle Scholar
  30. 30.
    Shi XJ, Yeung L, Lam WY, Wu PKS, Zhou BS (2009) Protein profiles in zebrafish (Danio rerio) embryos exposed to perfluorooctane sulfonate. Toxicol Sci 110:334–340PubMedCrossRefGoogle Scholar
  31. 31.
    Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
  32. 32.
    Garner DL (2006) Flow cytometric sexing of mammalian sperm. Theriogenology 65:943–957PubMedCrossRefGoogle Scholar
  33. 33.
    Howes EA, Miller NG, Dolby C, Hutchings A, Butcher GW, Jones R (1997) A search for sex-specific antigens on bovine spermatozoa using immunological and biochemical techniques to compare the protein profiles of X and Y chromosome-bearing sperm populations separated by fluorescence-activated cell sorting. J Reprod Fertil 110:195–204PubMedCrossRefGoogle Scholar
  34. 34.
    Soares S, Barbosa J (2008) Application of recombinant antibody library for screening specific antigens in a bovine sperm cell subpopulation. Livest Sci 114:188–193CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Wu-Cai Yang
    • 1
    • 2
  • Lei Sang
    • 3
  • Yao Xiao
    • 2
  • Hua-Lin Zhang
    • 2
  • Ke-Qiong Tang
    • 2
  • Li-Guo Yang
    • 2
  1. 1.College of Animal Science and TechnologyNorthwest A&F UniversityShaanxiChina
  2. 2.College of Animal Science and TechnologyHuazhong Agricultural UniversityWuhanChina
  3. 3.Institute of Animal Husbandry and Veterinary MedicineFujian Academy of Agricultural SciencesFuzhouChina

Personalised recommendations