Evaluation of Fertilizing Ability of Frozen Equine Sperm Using a Bovine Zona Pellucida Binding Assay.

The researchers have developed an assay to measure the fertility of frozen horse sperm using a bovine Zona Pellucida binding assay (ZBA). This method might help predict the viability of the frozen equine sperm better than existing methods.

Overview of the Study

• The study was prompted by the observation that frozen horse semen has lower fertility rates compared to freshly cooled semen.
• Collecting horse oocytes (eggs) is challenging, so an alternate method for predicting the fertility potential of frozen horse sperm was needed. The researchers decided to use a heterologous Zona Pellucida Binding Assay (ZBA), that uses cow eggs instead of horse eggs.
• Importantly, the study aimed to predict fertilizing ability by observing the rate of capacitated and hyperactivated sperm, which are specific stages needed for successful fertilization, with the help of a Computer Assisted Sperm Analyzer.

Methods & Procedures Used

• In order to evaluate the fertilizing ability of the frozen equine sperm, they applied a bovine (cow) ZBA. An in vitro capacitation and hyperactivation media with procaine and calcium ionophore A23187 were used, respectively.
• They calculated the sperm motility characteristics, rates of intact and acrosome-reacted sperm, and the number of the stallion sperm that bound to the bovine ZP (egg).

Results of the Study

• The group that received procaine showed a hyperactivation motility pattern, which is one of the factors important for successful fertilization.
• Despite this, procaine treatment improved the binding of sperm to the Zona Pellucida (ZP) similarly to how standard capacitation treatment does. Capacitation is another step in sperm preparation for successful fertilization.
• Interestingly, the capacitation medium boosted the In Vitro Fertilization (IVF) capability of frozen equine sperm, increasing the likelihood of successful sperm-oocyte (egg) interaction.

Conclusions of the Study

• The study concludes that the capacitation medium improves the ability of frozen equine sperm to fertilize an egg.
• This provides a promising route for maximizing the fertility of frozen equine sperm, which could have significant applications in horse breeding.
• It’s also the first time that in vitro hyperactivation and the heterologous ZBA methods have been used to evaluate the viability of frozen equine sperm.