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Reproduction & fertility2024; 5(4); e230081; doi: 10.1530/RAF-23-0081

Optimizing equine sperm quality: an alternative to single layer centrifugation for sperm isolation.

Abstract: In vitro semen purification techniques have been developed that seek to mimic the in vivo selection process in order to generate the highest possible chance of oocyte fertilization following artificial insemination. Numerous methods have been developed to isolate functional spermatozoa for artificial insemination, yet only one method, single-layer centrifugation using commercial preparations like EquiPure, has been widely employed. In this study, we have introduced a novel approach for isolating spermatozoa and compared their quality to those isolated using EquiPure. The AI port system (Memphasys, Ltd. in Sydney, Australia) features a disposable cartridge with an inoculation chamber for depositing extended semen and a harvest chamber for extracting isolated spermatozoa. These chambers are separated by a 5 µm polyethylene terephthalate (PETE) membrane, allowing highly motile spermatozoa to migrate from the inoculation chamber to the harvest chamber over a 20-minute period. This migration effectively leaves behind seminal plasma and other cell types, such as leukocytes. Comparative analyses between spermatozoa isolated with the AI port and EquiPure demonstrated that, across all measured sperm parameters, including yield, vitality, motility, morphology, DNA fragmentation, and mitochondrial superoxide generation, the AI port-isolated cells exhibited comparable or superior performance, particularly in terms of DNA fragmentation. In summary, the AI port system demonstrates the potential to efficiently isolate high-quality spermatozoa, possibly offering a cost-effective and user-friendly alternative that may enhance the success rates of artificial insemination in breeding programs. Unassigned: This study aimed to create a new method for refining stallion semen to increase the likelihood of a successful pregnancy through artificial insemination. While there are existing techniques for isolating high-quality sperm, the most common involves a complicated process using a centrifuge, which spins the semen to separate it. This research introduces a new approach called the AI port system that uses a disposable cartridge with two separate chambers for putting in semen and getting out isolated sperm. A membrane between the chambers acts like a filter, letting highly motile sperm swim across, leaving behind unwanted substances like bacteria and blood cells. Compared to the centrifugation method, the AI port system effectively produces sperm with comparable or better quality in various aspects, including vitality, movement, shape, DNA integrity, and energy production. In summary, the AI port system is an easy-to-use alternative with the potential to improve the success of artificial insemination in horse breeding programs.
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Publication Date: 2024-11-11 PubMed ID: 39437190PubMed Central: PMC11558927DOI: 10.1530/RAF-23-0081Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

Overview

  • This study introduces the AI port system, a novel sperm isolation method for horses, and compares its effectiveness to the widely used single-layer centrifugation technique (EquiPure) in improving sperm quality for artificial insemination.

Background and Motivation

  • In vitro semen purification techniques aim to select the best quality spermatozoa to increase the chances of fertilization.
  • Many sperm isolation methods exist, but single-layer centrifugation with commercial products like EquiPure is the most common for equines.
  • Centrifugation methods can be complex, equipment-intensive, and may require technical expertise.
  • There is a need for an alternative sperm isolation method that is simple, cost-effective, and capable of producing high-quality spermatozoa.

The AI Port System: Description and Function

  • Developed by Memphasys, Ltd., the AI port system uses a disposable cartridge with two distinct chambers:
    • An inoculation chamber where extended semen is deposited.
    • A harvest chamber from which isolated spermatozoa are collected.
  • The two chambers are separated by a 5 µm polyethylene terephthalate (PETE) membrane that acts as a selective barrier.
  • Highly motile spermatozoa swim through the membrane into the harvest chamber within 20 minutes.
  • Unwanted components such as seminal plasma, leukocytes, bacteria, and other cells remain behind in the inoculation chamber.
  • This process mimics natural sperm selection without the need for centrifugation.

Comparative Analysis of Sperm Quality

  • The study compared sperm isolated using the AI port system to sperm isolated using EquiPure single-layer centrifugation on various parameters:
    • Yield: The quantity of sperm recovered.
    • Vitality: The proportion of live sperm cells.
    • Motility: The ability of sperm to move actively and properly.
    • Morphology: The structural normality of sperm cells.
    • DNA Fragmentation: The integrity and breakage of sperm DNA, critical for fertility.
    • Mitochondrial Superoxide Generation: An indicator of sperm oxidative stress and health.
  • The AI port system demonstrated comparable or better results than EquiPure across all parameters, with significantly improved DNA fragmentation levels.

Advantages and Implications

  • The AI port system offers several potential benefits:
    • Simple and user-friendly operation without the need for centrifugation equipment.
    • Disposable cartridge design reduces contamination risk and maintenance requirements.
    • Cost-effective alternative for sperm isolation in breeding programs.
    • Enhanced sperm quality, particularly in genetic integrity, which may improve fertilization success and pregnancy rates.
  • This method may represent a practical improvement for artificial insemination protocols in equine breeding.
  • By better mimicking natural sperm selection processes, it could optimize reproductive outcomes while lowering technical barriers.

Conclusion

  • The AI port system is an innovative and efficient alternative to single-layer centrifugation for isolating high-quality stallion spermatozoa.
  • This technology has the potential to streamline artificial insemination practices and improve breeding success in the equine industry.
  • Further studies and field evaluations could solidify its role in routine use and uncover additional benefits.

Cite This Article

APA
Medica AJ, Gibb Z, Aitken RJ. (2024). Optimizing equine sperm quality: an alternative to single layer centrifugation for sperm isolation. Reprod Fertil, 5(4), e230081. https://doi.org/10.1530/RAF-23-0081

Publication

Reproduction & fertility
ISSN: 2633-8386
NlmUniqueID: 101778727
Country: England
Language: English
Volume: 5
Issue: 4
PII: e230081

Researcher Affiliations

Medica, Ashlee Jade
  • HMRI Infertility and Reproduction Research Program, Discipline of Biological Sciences, School of Environmental and Life Sciences, College of Engineering Science and Environment, University of Newcastle, Callaghan, New South Wales, Australia.
Gibb, Zamira
  • HMRI Infertility and Reproduction Research Program, Discipline of Biological Sciences, School of Environmental and Life Sciences, College of Engineering Science and Environment, University of Newcastle, Callaghan, New South Wales, Australia.
Aitken, Robert John
  • HMRI Infertility and Reproduction Research Program, Discipline of Biological Sciences, School of Environmental and Life Sciences, College of Engineering Science and Environment, University of Newcastle, Callaghan, New South Wales, Australia.

MeSH Terms

  • Animals
  • Male
  • Spermatozoa / cytology
  • Spermatozoa / physiology
  • Horses
  • Centrifugation / methods
  • Semen Analysis / methods
  • Semen Analysis / veterinary
  • Semen Analysis / instrumentation
  • Cell Separation / methods
  • Cell Separation / veterinary
  • Cell Separation / instrumentation
  • Sperm Motility / physiology
  • Insemination, Artificial / veterinary
  • Insemination, Artificial / methods
  • DNA Fragmentation

Conflict of Interest Statement

The financial support for the Ph.D. student who conducted these studies, AJ Medica, was granted through a scholarship awarded by Memphasys Ltd, a biotechnology company specializing in cell separation technologies.

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