Coulter counter-based evaluation of sperm volume to assess sperm viability of bull semen and application to X/Y sperm sorting

Abstract

The Coulter Counter Hypo-Osmotic Swelling test (CC-HOS) was developed to provide insight into the membrane integrity (relative volume shift Vr) of sperm necessary for fertilization, and to identify the optimum buffer needed for the X/Y chromosome sorting process. Using the CC-HOS test on neat bovine semen, the mean relative volume shift Vr for July and August was 1.20 and 1.14, respectively, whereas mean Vr values ranged from 1.32 to 1.41 during September to November. There was an inverse relationship between Vr magnitude and environmental temperature; we inferred that this enhanced sperm viability during autumn relative to summer. A method was developed to measure the dynamics of volume change of sperm in the buffer (pH 6.5) used for the X/Y chromosome sorting process. When exposed to the buffer (4 mM K+, 153 mM Na+, 140 mM Cl(-)), sperm from Bull C had a mean modal volume of 22.8+/-0.2 fL during a 0-300 s time interval, which did not significantly vary from sperm volumes (21.88+/-0.66 fL for Bull A and 22.46+/-0.38 fL for Bull B) noted in isotonic Isoton II solution. However, when exposed to lower ionic concentrations (2 mM K+, 62 mM Na+, 47 mM Cl-), the mean volume of Bull C sperm increased to 29.2+/-1.5 fL and exhibited slower rates toward stabilized volumes relative to higher ionic concentration buffers. Utilization of volume swelling measurements for measuring the impact of ion concentrations in X/Y chromosome sorting process buffers illustrated the importance of its application for emerging sperm-based biotechnologies.