PSXII-14 - Estimation of Phosphatidylserine positive sperm in fresh bull semen

Previous research demonstrated that phosphatidylserine (PS), a marker for apoptosis, plays a role in murine sperm fertilization capacity; however, less is known of the role of PS in bovine fertilization. The objective of this experiment was to determine the concentration of PS positive sperm on fresh bovine semen (n = 4). A breeding soundness evaluation was performed and electroejaculation was used to collect semen which was immediately evaluated for ejaculate density and gross motility through visual evaluation. Semen smears of semen mixed with an eosin-nigrosin strain were prepared on glass microscope slides. Semen was mixed with OptiXcell™ (IMV Technologies) semen extender, a 0.5 mL sample from each bull was washed with non-capacitating media and then incubated at 37.5°C 5% CO₂ with capacitating media for 60 minutes. Samples were stained with Annexin V FITC and 7AAD for 15 minutes each. Samples were analyzed using a Flow Cytometer to determine the percentage of PS positive sperm. Bull 1 had a scrotal circumference of 37.5 cm, ejaculate density of 400-750 million sperm/mL, very good gross motility, 78% sperm with normal morphology, and 14.65% PS positive sperm. Bull 2 had a scrotal circumference of 37 cm, ejaculate density of 400-750 million sperm/mL, very good gross motility, 84% sperm with normal morphology, and 17.05% PS positive sperm. Bull 3 had a scrotal circumference of 39.5 cm, ejaculate density of 250-400 million sperm/mL, fair gross motility, 48% sperm with normal morphology, and 12.8% PS positive sperm. Bull 4 had a scrotal circumference of 38 cm, ejaculate density of 400-750 million sperm/mL, good gross motility, 68% of sperm with normal morphology, and 12.65% PS positive sperm. Our data demonstrate the possibility of identifying live PS positive sperm in fresh ejaculate from bulls. The possibility exists that PS plays a role in sperm oocyte fusion and fertilization in the bovine.