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Optimization of microelectrophoresis to select highly negatively charged sperm

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Abstract

Purpose

The sperm membrane undergoes extensive surface remodeling as it matures in the epididymis. During this process, the sperm is encapsulated in an extensive glycocalyx layer, which provides the membrane with its characteristic negative electrostatic charge. In this study, we develop a method of microelectrophoresis and standardize the protocol to isolate sperm with high negative membrane charge.

Methods

Under an electric field, the percentage of positively charged sperm (PCS), negatively charged sperm (NCS), and neutrally charged sperm was determined for each ejaculate prior to and following density gradient centrifugation (DGC), and evaluated for sperm DNA damage, and histone retention. Subsequently, PCS, NCS, and neutrally charged sperm were selected using an ICSI needle and directly analyzed for DNA damage.

Results

When raw semen was analyzed using microelectrophoresis, 94 % were NCS. In contrast, DGC completely or partially stripped the negative membrane charge from sperm resulting PCS and neutrally charged sperm, while the charged sperm populations are increased with an increase in electrophoretic current. Following DGC, high sperm DNA damage and abnormal histone retention were inversely correlated with percentage NCS and directly correlated with percentage PCS. NCS exhibited significantly lower DNA damage when compared with control (P < 0.05) and PCS (P < 0.05). When the charged sperm population was corrected for neutrally charged sperm, sperm DNA damage was strongly associated with NCS at a lower electrophoretic current.

Conclusion

The results suggest that selection of NCS at lower current may be an important biomarker to select healthy sperm for assisted reproductive treatment.

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Acknowledgments

This project was supported by research grants from EMD Serono, Rockland, Massachusetts, and the Howard and Georgeanna Jones Foundation for Reproductive Medicine, Norfolk, Virginia. The authors wish to thank the UCRM IVF unit and laboratory staff for their commitment and support to this project, for preparing tissue samples, and for helping to collect data on ART outcomes.

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Authors and Affiliations

  1. Andrology and IVF Laboratory, Department of Surgery (Urology), Salt Lake City, Utah, USA

    Luke Simon, Kristin Murphy, Kenneth I. Aston, Benjamin R. Emery, James M. Hotaling & Douglas T. Carrell

  2. Department of Obstetrics and Gynecology, Salt Lake City, Utah, USA

    Douglas T. Carrell

  3. Department of Human Genetics, University of Utah, 675 Arapeen Drive, Suite 201, Salt Lake City, Utah, 84109, USA

    Douglas T. Carrell

Authors
  1. Luke Simon
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  2. Kristin Murphy
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  3. Kenneth I. Aston
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  4. Benjamin R. Emery
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  5. James M. Hotaling
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  6. Douglas T. Carrell
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Corresponding author

Correspondence to Douglas T. Carrell.

Additional information

Capsule This study is aimed to standardize the microelectrophoresis protocol to isolate sperm with high negative membrane charge. The results from this study suggest that selection of negatively charged sperm at lower current may serve as an important biomarker to select healthy sperm for assisted reproductive treatment.

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Simon, L., Murphy, K., Aston, K.I. et al. Optimization of microelectrophoresis to select highly negatively charged sperm. J Assist Reprod Genet 33, 679–688 (2016). https://doi.org/10.1007/s10815-016-0700-x

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  • DOI: https://doi.org/10.1007/s10815-016-0700-x

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