[Single- and two-layer gradient centrifugation in sperm separation: comparison and appraisal]

Area of Science:

• Reproductive biology and andrology
• Sperm preparation techniques

Background:

• Sperm separation is crucial for assisted reproductive technologies.
• Percoll density gradient centrifugation is a common method for sperm preparation.
• Evaluating different Percoll gradient configurations is essential for optimizing sperm yield and quality.

Purpose of the Study:

• To compare the efficacy of single-layer versus two-layer Percoll density gradient centrifugation for sperm separation.
• To assess the impact of each method on sperm recovery rate, motility parameters, and round cell depletion.

Main Methods:

• Twenty human semen specimens were processed using both single-layer (50% Percoll) and two-layer (90% and 45% Percoll) density gradient centrifugation.
• Sperm Class Analyzer (SCA) was employed to evaluate sperm density, motility (grades a, b, c), dynamic parameters, and round cell density pre- and post-treatment.
• Statistical analysis was performed to compare the outcomes of the two methods.

Main Results:

• The single-layer method demonstrated a significantly higher sperm recovery rate (65.5 ± 12.8%) compared to the two-layer method (P < 0.01).
• Both methods significantly increased the percentage of grade 'a' sperm post-treatment (P < 0.05, P < 0.01). The two-layer method showed a significant increase in grade 'a+b' sperm (P < 0.05), while the single-layer method did not (P > 0.05).
• Both techniques effectively reduced round cell density (P < 0.05, P < 0.01) without significant differences between them.

Conclusions:

• Single-layer Percoll centrifugation provides superior sperm recovery with minimal impact on sperm motility.
• Two-layer Percoll centrifugation results in lower sperm recovery but significantly enhances sperm motility parameters.
• Both methods are effective in removing round cells, offering distinct advantages for specific in vitro applications.