Effect of density-gradient sperm separation technique on in vitro fertilization potential of water buffalo semen with low post-thaw motilities

The use of discontinuous colloidal suspension of silica particles were tested to separate the live spermatozoa from seminal constituents to improve the in vitro fertilization (IVF) potential of semen with low post-thaw sperm motility. Three different suspensions combined to form discontinuous gradients: 45% and 95% (45/95),65% and 95% (65/95), and 45%,65% and 95% (45/65/95) were examined to compare separation efficiency with semen prepared by the conventional method which served as the control. In vitro matured oocytes were used for IVF. Recovery and concentration of motile sperm cells, embryo cleavage and development and Improvement in sperm motility between bulls were assessed. Density gradient successfully separated the motile spermatozoa increasing sperm motility for fertilization from 23.90 +/- 1.23% to 96.40 +/- 1.9% against 25.65 +/- 4.0% in the control group. Combination of three discontinuous layers (45/65/95) had significantly (P<0.01) purified motile sperms. Sperm concentration decreased from 41.85 +/- 0.44 M to 9.00 +/- 4.9M with purely motile spermatozoa. The separated spermatozoa for IVF led to significantly (P<0.01) improved cleavage (69.1 +/- 2.04%) and embryo development (23.6 +/- 1.0%) rates. Significant differences (P<0.01) on sperm motility exist among bulls. Density gradient sperm separation improved the sperm motility of all bulls indicating potential use of the technique in improving the reproductive potential of sperm with low post-thaw motility.