Silica-based colloid centrifugation enhances sperm quality in cockerel semen

Abstract

1. Percoll^TM is one of the most widely used colloid for animal sperm preparation. The aim of this study was to evaluate whether Percoll^TM colloid centrifugation could be practical to improve cockerel sperm quality, and to compare the effects of Percoll^TM single layer centrifugation (SLC) and density gradient centrifugation (DGC) in order to obtain the most optimal protocol for cockerel semen.2. In the experiment with Percoll^TM SLC for fresh semen, an increase of motile sperm was seen after Percoll^TM 80% SLC and 90% SLC was conducted, at levels of 28.8% and 30.2% respectively (P < 0.01). The increase of progressively motile sperm after Percoll^TM 80% SLC and 90% SLC was 177.2% and 202.4% respectively (P < 0.01). Meanwhile, for semen stored at 4°C for 24 h, the increase of motile sperm after Percoll^TM 70% SLC and 80% SLC was 41.2% and 44.0% (P < 0.01), and the increase of progressive sperm after Percoll^TM 70% SLC and 80% SLC was 71.3% and 83.1% respectively (P < 0.01). Both the percentage of motile sperm and progressive sperm of the fresh and stored cockerel semen after appropriate Percoll^TM SLC was significantly enhanced.3. Sperm membrane integrity did not show any decrease after Percoll^TM centrifugation compared with non-centrifuged semen, which suggested that the Percoll^TM centrifugation treatment in this study did not cause damage to cockerel sperm membranes.4. In the experiment regarding the comparison of Percoll^TM SLC and DGC with fresh semen, the increase of motile sperm after Percoll^TM 80% SLC, 90% SLC and 40%/80% DGC was 29.5%, 36.4%, and 25.0% respectively; and the increase of progressive sperm was 44.7%, 58.5%, and 54.7%, respectively. For semen stored at 4°C for 24 h, the increase of motile sperm after Percoll^TM 70% SLC, 80% SLC and 35%/70% DGC were 41.2%, 44.0%, and 26.4%; and the increase of progressive sperm was 71.3%, 83.1%, and 43.7%, respectively. There were no significant differences between the increase of sperm motility after Percoll^TM 80%, 90% SLC or Percoll^TM 40%/80% DGC in fresh cockerel semen. There was no significant difference between Percoll^TM 70%, 80% SLC and Percoll^TM 35%/70% in stored cockerel semen. There was a tendency for sperm recovery rates with Percoll^TM SLC to be higher than Percoll^TM DGC, although this did not reach statistical significance in this study.5. It was concluded that Percoll^TM SLC was more suitable for cockerel sperm separation than Percoll^TM DGC. The results suggested that Percoll^TM 80% SLC was the most optimal procedure to separate fresh cockerel sperm and Percoll^TM 70% SLC was the most optimal procedure to separate stored cockerel sperm. Percoll^TM SLC is more simple, user-friendly and economical and less time-consuming than DGC for cockerel semen processing.

Keywords: CASA; PercollTM; density gradient centrifugation (DGC); flow cytometry; membrane integrity; motility; single layer centrifugation (SLC).