Sperm-binding to the perivitelline membrane of chicken egg yolk as a functional test for sperm evaluation in dogs

Abstract

During fertilization, spermatozoa interact with the zona pellucida (ZP) through the binding between the acrosome and proteins 2 and 3 (ZP2 and ZP3). The perivitelline membrane of chicken egg yolk is homologous to the mammalian ZP3, which allows the binding of sperm of several species. The aim of this study was to standardize and evaluate the efficiency of sperm-binding to the perivitelline membrane of chicken eggs as a functional method for canine semen evaluation. For this purpose, nine post-thaw sperm samples were used, which were divided into two aliquots: the first was kept in water bath at 37ºC (live sample) and the second was submitted to cold shock to induce cellular damage (dead sample). The two aliquots were mixed on five proportions, corresponding to 0%, 25%, 50%, 75%, and 100% of viable cells, and the binding test was performed by analyzing the number of spermatozoa bonded to the perivitelline membrane by means of computerized assessment of sperm motility (CASA) or conventional microscopy. Additionally, samples were submitted to sperm motility analysis, evaluation of plasmatic and acrosomal membrane integrity, and sperm mitochondrial activity. The sperm-binding test to the perivitelline membrane of chicken egg yolk was considered a feasible sperm analysis test for both fertilizing capacity and overall sperm attributes evaluation, mainly when the analysis is performed by a conventional microscope, which expands its practicality to the majority of canine reproduction laboratories.

Keywords

• Sperm cryopreservation
• Egg yolk
• Sperm analysis
• Perivitelline membrane
• Canine