Optimizing human semen cryopreservation by reducing test vial volume and repetitive test vial sampling

Area of Science:

• Reproductive Medicine
• Cryobiology
• Andrology

Background:

• Optimizing semen cryopreservation is crucial for assisted reproductive technologies (ART).
• Standardization of test vial (TV) volumes and storage conditions is needed for reliable cryosurvival (CS) prediction.

Purpose of the Study:

• To determine the optimal test vial (TV) volume for human semen cryopreservation.
• To evaluate the reliability of TVs for predicting cryosurvival (CS).
• To assess the impact of intermediate temperature exposure and long-term storage in nitrogen vapor (VN2) versus liquid nitrogen (LN2) on semen viability.

Main Methods:

• A prospective clinical laboratory study involving 594 patients undergoing semen analysis and cryopreservation.
• Evaluation of various test vial (TV) volumes (50-1,000 μL) and cryostorage conditions, including intermediate temperatures (-88°C to -93°C) and storage in VN2 and LN2.
• Analysis of post-thaw motility and cryosurvival (CS) rates.

Main Results:

• A 50 μL TV volume resulted in lower cryosurvival (CS) compared to larger volumes; 100 μL TVs showed similar CS to larger volumes.
• Intermediate temperature exposure did not negatively impact post-thaw motility.
• Cryosurvival (CS) in test vials (TVs) and ART procedure vials (ARTVs) from the same ejaculate were comparable, with strong correlation between the first TV and subsequent ARTVs.
• Long-term cryostorage in nitrogen vapor (VN2) did not affect cryosurvival (CS).

Conclusions:

• A single 100 μL TV per patient is sufficient to predict cryosurvival (CS) when freezing multiple ejaculates within a short timeframe (<10 days).
• Semen cryostorage in nitrogen vapor (VN2) offers a stable and safe long-term preservation environment.