Effects of centrifugation, glycerol level, cooling to 5 degrees C, freezing rate and thawing rate on the post-thaw motility of equine sperm.

This study examines the impact of various processing, freezing, and thawing techniques on the motility of equine sperm after thawing. The researchers identified a new procedure that improves post-thaw motility of equine sperm by 12% compared to previous methods.

Methods of the Study

• The study involved conducting five separate experiments to evaluate the effects of different processing strategies—including centrifugation, varying levels of glycerol, cooling rates, freezing rates, and thawing rates—on the post-thaw motility of horse sperm.
• The centrifugation process involved diluting the semen with a citrate-based medium before centrifuging. The result showed that dilution to 50 x 10(6) sperm/ml with a centrifugation speed of 400 x g for 15 minutes yielded the best sperm motility.
• The inclusion of glycerol—a common cryoprotectant in sperm freezing—was also examined. The researchers found that using a 4% glycerol extender resulted in better post-thaw motility than either 2% or 6% glycerol.

Results and Conclusions

• Two different rates of cooling were compared: a slow cooling rate from 20 degrees C to -120 degrees C via 5 degrees C and a continuous cooling from 20 degrees C to -120 degrees C. No significant differences in sperm motility were observed between the two cooling techniques.
• Sperm thawing rates were evaluated, with thawing in 75 degrees C water for 7 seconds proving superior to thawing in 37 degrees C water for 30 seconds.
• Based on these experiments, the researchers established a more effective procedure for processing, freezing, and thawing semen. This involves dilution of semen in a specific centrifugation medium, resuspension in a lactose-EDTA-egg yolk extender with 4% glycerol, packaging, specific freezing rates, and a fast thawing method.
• With this new procedure, the post-thaw motility of sperm improved by 12%, up from 22% using previous methods to 34%.

Significance of the Study

• The findings of this study are particularly important for equine breeding and fertility practices, as the new procedure offers a significant improvement in post-thaw horse sperm motility.
• This research underlines the importance of careful handling during sperm processing, especially when it comes to centrifugation, the use of glycerol, cooling rates, freezing rates, and thawing rates. It shows that each of these factors can have a significant influence on post-thaw sperm motility.