Skip to main content
SpringerLink
Log in

Femtosecond laser is effective tool for zona pellucida engraving and tagging of preimplantation mammalian embryos

  • Technological Innovations
  • Published:
Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

Our purpose was to study whether application of femtosecond laser pulses for alphanumeric code marking in the volume of zona pellucida (ZP) could be effective and reliable approach for direct tagging of preimplantation embryos.

Methods

Femtosecond laser pulses (wavelength of 514 nm, pulse duration of 280 fs, repetition rate of 2.5 kHz, pulse energy of 20 nJ) were applied for precise alphanumeric code engraving on the ZP of mouse embryos at the zygote stage for individual embryo marking and their accurate identification. Embryo quality assessment every 24 h post laser-assisted marking as well as immunofluorescence staining (for ICM/TE cell number ratio calculation) were performed.

Results

Initial experiments have started with embryo marking in a single equatorial plane. The codes engraved could be clearly recognized until the thinning of the ZP prior to hatching. Since embryo may change its orientation during the ART cycle, multi-plane code engraving seems to be more practical for simplifying the process of code searching and embryo identification. We have marked the ZP in three planes, and no decrease in developmental rates as well as no morphological changes of embryos post laser-assisted engraving have been observed as compared to control group embryos.

Conclusions

Our results demonstrate the suitability of femtosecond laser as a novel tool for noninvasive embryo tagging, enabling embryo identification from day 0.5 post coitum to at least early blastocyst stage. Thus, the versatility and the potential use of femtosecond lasers in the field of developmental biology and assisted reproduction have been shown.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Bedient C, Khanna P, Desai N. Laser pulse application in IVF. In: Jakubczak K, editor. Lasers - applications in science and industry: InTech; 2011. p. 193–214.

  2. Karu TI. Lasers in infertility treatment: irradiation of oocytes and spermatozoa. Photomed Laser Surg. 2012;30:239–41. https://doi.org/10.1089/pho.2012.9888.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Abdel-Salam Z, Harith MA. Laser researches on livestock semen and oocytes: a brief review. J Adv Res. 2015;6:311–7. https://doi.org/10.1016/j.jare.2014.11.006.

    Article  CAS  PubMed  Google Scholar 

  4. Montag M, Rink K, Delacretaz G, van der Ven H. Laser induced immobilization and plasma membrane permeabilization in human spermatozoa. Hum Reprod. 2000;15:846–52. https://doi.org/10.1093/humrep/15.4.846.

    Article  CAS  PubMed  Google Scholar 

  5. Sato H, Landthaler M, Haina D, Schill WB. The effects of laser light on sperm motility and velocity in vitro. Andrologia. 1984;16:23–5.

    Article  CAS  PubMed  Google Scholar 

  6. Lenzi A, Claroni F, Gandini L, Lombardo F, Barbieri C, Lino A, et al. Laser radiation and motility patterns of human sperm. Syst Biol Reprod Med. 1989;23:229–34.

    CAS  Google Scholar 

  7. Preece D, Chow KW, Gomez-Godinez V, Gustafson K, Esener S, Ravida N, et al. Red light improves spermatozoa motility and does not induce oxidative DNA damage. Sci Rep. 2017;7:46480. https://doi.org/10.1038/srep46480.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Zhang H, Liu KK. Optical tweezers for single cells. J R Soc Interface. 2008;5:671–90. https://doi.org/10.1098/rsif.2008.0052.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Nascimento JM, Shi LZ, Meyers S, Gagneux P, Loskutoff NM, Botvinick EL, et al. The use of optical tweezers to study sperm competition and motility in primates. J R Soc Interface. 2008;5:297–302. https://doi.org/10.1098/rsif.2007.1118.

    Article  PubMed  Google Scholar 

  10. Clement-Sengewald A, Schütze K, Ashkin A, Palma GA, Kerlen G, Brem G. Fertilization of bovine oocytes induced solely with combined laser microbeam and optical tweezers. J Assist Reprod Genet. 1996;13:259–65. https://doi.org/10.1007/BF02065947.

    Article  CAS  PubMed  Google Scholar 

  11. Clement-Sengewald A, Buchholz T, Schütze K, Berg U, Berg FD. Noncontact, laser-mediated extraction of polar bodies for prefertilization genetic diagnosis. J Assist Reprod Genet. 2002;19:183–94. https://doi.org/10.1023/A:1014894029099.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ilina IV, Rakityanskiy MM, Sitnikov DS, Ovchinnikov AV, Agranat MB, Khramova YV, et al. Biomedical and biotechnology applications of noncontact femtosecond laser microsurgery of living cells. AIP Conf Proc. 2012;1464:560–71. https://doi.org/10.1063/1.4739909.

    Article  Google Scholar 

  13. Douglas-Hamilton DH, Conia J. Thermal effects in laser-assisted pre-embryo zona drilling. J Biomed Opt. 2001;6:205–13.

    Article  CAS  PubMed  Google Scholar 

  14. Tucker MJ, Ball GD. Assisted hatching as a technique for use in human in vitro fertilization and embryo transfer is long overdue for careful and appropriate study. J Clin Embr. 2009;12:5–8.

    Google Scholar 

  15. Taylor T, Gilchrist J, Hallowell S, Hanshew K, Orris J, Glassner M, et al. The effects of different laser pulse lengths on the embryo biopsy procedure and embryo development to the blastocyst stage. J Assist Reprod Genet. 2010;27:663–7. https://doi.org/10.1007/s10815-010-9461-0.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Stevenson DJ, Gunn-Moore FJ, Campbell P, Dholakia K. Single cell optical transfection. J R Soc Interface. 2010;7:863–71. https://doi.org/10.1098/rsif.2009.0463.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Torres-Mapa ML, Antkowiak M, Cizmarova H, Ferrier DEK, Dholakia K, Gunn-Moore FJ. Integrated holographic system for all-optical manipulation of developing embryos. Biomed Opt Express. 2011;2:1564–75. https://doi.org/10.1364/BOE.2.001564.

    Article  PubMed  PubMed Central  Google Scholar 

  18. Ilina IV, Ovchinnikov AV, Sitnikov DS, Rakityanskiy MM, Agranat MB, Khramova YV, et al. Application of femtosecond laser pulses in biomedical cell technologies. High Temp. 2013;51:173–8. https://doi.org/10.1134/S0018151X13020089.

    Article  CAS  Google Scholar 

  19. Osychenko AA, Zalessky AD, Krivokharchenko AS, Shakhbazian AK, Ryabova AV, Nadtochenko VA. Fusion of blastomeres in mouse embryos under the action of femtosecond laser radiation. Efficiency of blastocyst formation and embryo development. Quantum Electron. 2015;45:498–502. https://doi.org/10.1070/QE2015v045n05ABEH015767.

    Article  CAS  Google Scholar 

  20. Kuetemeyer K, Lucas-Hahn A, Petersen B, Lemme E, Hassel P, Niemann H, et al. Combined multiphoton imaging and automated functional enucleation of porcine oocytes using femtosecond laser pulses. J Biomed Opt. 2010;15:046006. https://doi.org/10.1117/1.3463012.

    Article  PubMed  Google Scholar 

  21. Ilina IV, Khramova YV, Filatov MA, Semenova ML, Sitnikov DS. Application of femtosecond laser scalpel and optical tweezers for noncontact biopsy of late preimplantation embryos. High Temp. 2015;53:804–9. https://doi.org/10.1134/S0018151X15060103.

    Article  CAS  Google Scholar 

  22. Liebler R. Are you my parent? Are you my child? The role of genetics and race in defining relationships after reproductive technological mistakes. DePaul J Health Care L. 2002;5:15–56.

    Google Scholar 

  23. Spriggs M. 2003 IVF mixup: white couple have black babies. J Med Ethics. 2003;29:65. https://doi.org/10.1136/jme.29.2.65.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Bender L. ‘To err is human’. ART mix-ups: a labor-based, relational proposal. J Gender Race & Just. 2006;9:1–90.

    Google Scholar 

  25. Forte M, Faustini F, Maggiulli R, Scarica C, Romano S, Ottolini C, et al. 2016 electronic witness system in IVF—patients perspective. J Assist Reprod Genet. 2016;33:1215–22. https://doi.org/10.1007/s10815-016-0759-4.

    Article  PubMed  PubMed Central  Google Scholar 

  26. de los Santos MJ, Ruiz A. Protocols for tracking and witnessing samples and patients in assisted reproductive technology. Fertil Steril. 2013;100:1499–502. https://doi.org/10.1016/j.fertnstert.2013.09.029.

    Article  PubMed  Google Scholar 

  27. The Practice Committe of the American Society for Reproductive Medicine and the Practice Committe of the Society for Assisted Reproductive Technology. Revised guidelines for human embryology and andrology laboratories. Fertil Steril. 2008;90:S45–59. https://doi.org/10.1016/j.fertnstert.2008.08.099.

    Article  Google Scholar 

  28. Glew AM, Hoha K, Graves J, Lawrence H, Read S, Ah-Moye M. Radio frequency identity tags ‘RFID’ for electronic witnessing of IVF laboratory procedures. Fertil Steril. 2006;86:S170. https://doi.org/10.1016/j.fertnstert.2006.07.454.

    Article  Google Scholar 

  29. Thornhill AR, Brunetti XO, Bird S. Measuring human error in the IVF laboratory using an electronic witnessing system // Proc. of 17th World Congress on Controversies in Obstetrics, Gynecology & Infertility (COGI). 2013;101–106.

  30. Schnauffer K, Kingsland C, Troup S. Barcode labelling in the IVF laboratory. Hum Reprod. 2005;20(suppl.1):i79–80.

    Google Scholar 

  31. Novo S, Nogues C, Penon O, Barrios L, Santalo J, Gomez-Martinez R, et al. Barcode tagging of human oocytes and embryos to prevent mix-ups in assisted reproduction technologies. Hum Reprod. 2014;29:18–28. https://doi.org/10.1093/humrep/det409.

    Article  PubMed  Google Scholar 

  32. Hur YS, Ryu EK, Park SJ, Yoon J, Yoon SH, Yang GD, et al. Development of a security system for assisted reproductive technology (ART). J Assist Reprod Genet. 2014;32:155–68. https://doi.org/10.1007/s10815-014-0367-0.

    Article  PubMed  PubMed Central  Google Scholar 

  33. Hogan B, Beddington R, Costantini F, Lacy E. Manipulating the mouse embryo: a laboratory manual. New York: Cold Spring Harbor Lab; 2014.

    Google Scholar 

  34. Yan Z, Liang H, Deng L, Long H, Chen H, Chai W, et al. Eight-shaped hatching increases the risk of inner cell mass splitting in extended mouse embryo culture. PLoS One. 2015;10:e0145172. https://doi.org/10.1371/journal.pone.0145172.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Germond M, Nocera D, Senn A, Rink K, Delacrétaz G, Fakan S. Microdissection of mouse and human zona pellucida using a 1.48-microns diode laser beam: efficacy and safety of the procedure. Fertil Steril. 1995;64:604–11.

    Article  CAS  PubMed  Google Scholar 

  36. Hsieh YY, Huang CC, Cheng TC, Chang CC, Tsai HD, Lee MS. Laser-assisted hatching of embryos is better than the chemical method for enhancing the pregnancy rate in women with advanced age. Fertil Steril. 2002;78:179–82.

    Article  PubMed  Google Scholar 

  37. Malter HE, Schimmel T, Cohen J. Zona dissection by infrared laser: developmental consequences in the mouse, technical considerations, and controlled clinical trial. Reprod BioMed Online. 2001;3:117–23.

    Article  PubMed  Google Scholar 

  38. Fedele D, Fusi F. Thermal effects of NIR laser radiation in biological tissue: a brief survey. Energy for Health. 2010;6:10–5.

    Google Scholar 

  39. Sagoskin AW, Han T, Graham JR, Levy MJ, Stillman RJ, Tucker MJ. Healthy twin delivery after day 7 blastocyst transfer coupled with assisted hatching. Fertil Steril. 2002;77:615–7.

    Article  PubMed  Google Scholar 

  40. Li MW, Kinchen KL, Vallelunga JM, Young DL, Wright KDK, Gorano LN, et al. Safety, efficacy and efficiency of laser-assisted IVF in subfertile mutant mouse strains. Reproduction. 2013;145:245–54. https://doi.org/10.1530/REP-12-0477.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Anzai M, Nishiwaki M, Yanagi M, Nakashima T, Kaneko T, Taguchi Y, et al. Application of laser-assisted zona drilling to in vitro fertilization of cryopreserved mouse oocytes with spermatozoa from a subfertile transgenic mouse. J Reprod Dev. 2006;52:601–6.

    Article  PubMed  Google Scholar 

  42. Karmenyan AV, Shakhbazyan AK, Sviridova-Chailakhyan TA, Krivokharchenko AS, Chiou AE, Chailakhyan LM. Use of picosecond infrared laser for micromanipulation of early mammalian embryos. Mol Reprod Dev. 2009;76:975–83. https://doi.org/10.1002/mrd.21045.

    Article  CAS  PubMed  Google Scholar 

  43. Vogel A, Noack J, Huttman G, Paltauf G. Mechanisms of femtosecond laser nanosurgery of cells and tissues. Appl Phys B Lasers Opt. 2005;81:1015–47. https://doi.org/10.1007/s00340-005-2036-6.

    Article  CAS  Google Scholar 

  44. Loesel FH, Fischer JP, Götz MH, Horvath C, Juhasz T, Noack F, et al. Non-thermal ablation of neural tissue with femtosecond laser pulses. Appl Phys B Lasers Opt. 1998;66:121–8.

    CAS  Google Scholar 

  45. Suhm N, Gotz MH, Fischer JP, Loesel F, Schlegel W, Sturm V, et al. Ablation of neural tissue by short-pulsed lasers–a technical report. Acta Neurochir. 1996;138:346–9. https://doi.org/10.1007/BF01411747.

    Article  CAS  PubMed  Google Scholar 

  46. Juhasz T, Kastis GA, Suarez C, Bor Z, Bron WE. Time-resolved observations of shock waves and cavitation bubbles generated by femtosecond laser pulses in corneal tissue and water. Lasers Surg Med. 1996;19:23–31. https://doi.org/10.1002/(SICI)1096-9101(1996)19:1<23::AID-LSM4>3.0.CO;2-S.

    Article  CAS  PubMed  Google Scholar 

  47. Oraevsky AA, Da Silva LB, Rubenchik AM, Feit MD, Glinsky ME, Perry MD, et al. 1996 plasma mediated ablation of biological tissues with nanosecond-to-femtosecond laser pulses: relative role of linear and nonlinear absorption. IEEE J Sel Top Quantum Electron. 1996;2:801–9. https://doi.org/10.1109/2944.577302.

    Article  CAS  Google Scholar 

  48. Feit MD, Rubenchik AM, Kim BM, da Silva LB, Perry MD. Physical characterization of ultrashort laser pulse drilling of biological tissue. Appl Surf Sci. 1998;127–129:869–74. https://doi.org/10.1016/S0169-4332(97)00758-7.

    Article  Google Scholar 

  49. Schwab B, Hagner D, Muller W, Lubatschowski H, Lenarz T, Heermann R. Bone ablation using ultrashort laser pulses. A new technique for middle ear surgery. Laryngorhinootologie. 2004;83:219–25. https://doi.org/10.1055/s-2004-814270.

    Article  CAS  PubMed  Google Scholar 

  50. Emigh B, An R, Hsu EM, Crawford TH, Haugen HK, Wohl GR, et al. Porcine cortical bone ablation by ultrashort pulsed laser irradiation. J Biomed Opt. 2012;17:028001. https://doi.org/10.1117/1.JBO.17.2.028001.

    Article  PubMed  Google Scholar 

  51. Jiang F, Yang X, Dai N, Lu P, Long H, Cui L. An in vitro study of femtosecond laser photodisruption in rabbit sclera. Front Optoelectron China. 2008;1:162–7. https://doi.org/10.1007/s12200-008-0022-4.

    Article  Google Scholar 

  52. Frederickson KS, White WE, Wheeland RG, Slaughter DR. Precise ablation of skin with reduced collateral damage using the femtosecond-pulsed, terawatt titanium-sapphire laser. Arch Dermatol. 1993;129:989–93.

    Article  CAS  PubMed  Google Scholar 

  53. Mian SI, Shtein RM. Femtosecond laser-assisted corneal surgery. Curr Opin Ophthalmol. 2007;18:295–9. https://doi.org/10.1097/ICU.0b013e3281a4776c.

    Article  PubMed  Google Scholar 

  54. Farid M, Steinert RF. Femtosecond laser-assisted corneal surgery. Curr Opin Ophthalmol. 2010;21:288–92. https://doi.org/10.1097/ICU.0b013e32833a8dbc.

    Article  PubMed  Google Scholar 

  55. Coskun S, Hollanders J, Al-Hassan S, Al-Sufyan H, Al-Mayman H, Jaroudi K. Day 5 versus day 3 embryo transfer: a controlled randomized trial. Hum Reprod. 2000;15:1947–52. https://doi.org/10.1093/humrep/15.9.1947.

    Article  CAS  PubMed  Google Scholar 

  56. Bungum M, Bungum L, Humaidan P, Yding AC. Day 3 versus day 5 embryo transfer: a prospective randomized study. Reprod BioMed Online. 2003;7:98–104.

    Article  CAS  PubMed  Google Scholar 

  57. Hatirnaz S, Perkas MK. Day 3 embryo transfer versus day 5 blastocyst transfers: a prospective randomized controlled trial. Turk J Obstet Gynecol. 2017;14:82–8. https://doi.org/10.4274/tjod.99076.

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The work was conducted using Unique Facility “Terawatt Femtosecond Laser Complex” in the Center for Collective Usage “Femtosecond Laser Complex” of JIHT RAS.

Funding

The reported study was funded by RFBR and Moscow city Government according to the research project no.19-32-70036.

Author information

Authors and Affiliations

  1. Joint Institute for High Temperatures of the Russian Academy of Sciences, Izhorskaya st. 13 Bd.2, Moscow, Russian Federation, 125412

    Inna V. Ilina & Dmitry S. Sitnikov

  2. Faculty of Biology, Lomonosov Moscow State University, Leninskie Gory, 12-1, Moscow, Russian Federation, 119234

    Yulia V. Khramova & Maxim A. Filatov

Authors
  1. Inna V. Ilina
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yulia V. Khramova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Maxim A. Filatov
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Dmitry S. Sitnikov
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Inna V. Ilina.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in the study involving animals were in accordance with the ethical standards of the Moscow State University.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ilina, I.V., Khramova, Y.V., Filatov, M.A. et al. Femtosecond laser is effective tool for zona pellucida engraving and tagging of preimplantation mammalian embryos. J Assist Reprod Genet 36, 1251–1261 (2019). https://doi.org/10.1007/s10815-019-01424-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10815-019-01424-x

Keywords

Search

Navigation