Optimized cryopreservation of mouse sperm based on fertilization rate

Area of Science:

• Reproductive Biology
• Cryobiology
• Mammalian Genetics

Background:

• Established in vitro fertilization (IVF) protocols exist for cryopreserved sperm.
• However, optimal cryoprotectant and cryopreservation methods for mouse sperm are not well-defined.
• This limits the efficiency of assisted reproductive technologies (ART) and sperm banking.

Purpose of the Study:

• To optimize cryopreservation protocols for C57BL/6J mouse sperm.
• To determine optimal raffinose concentration, cryoprotectant volume, and freezing conditions.
• To evaluate the impact of cryopreservation on sperm fertility and subsequent offspring viability.

Main Methods:

• In vitro fertilization (IVF) assays were used to assess sperm fertility.
• Sperm was cryopreserved using varying concentrations of raffinose in water as the cryoprotectant.
• Optimization of cryoprotectant volume and freezing height above liquid nitrogen was performed.
• Fertilized embryos were transferred to recipient mice to assess development into viable offspring.

Main Results:

• The optimal raffinose concentration was determined to be 23-24% (510-540 mOsm/kg).
• An optimal cryoprotectant volume of 180-400 µl per mouse was identified.
• Maximum sperm fertility was achieved when frozen 13-25 mm above liquid nitrogen.
• Raffinose in fertilization medium did not impede fertilization, and offspring were successfully produced.

Conclusions:

• Optimized cryopreservation protocols using raffinose significantly enhance mouse sperm fertility.
• These refined methods improve the efficiency of ART and sperm cryobanking.
• Successful development of viable offspring confirms the efficacy of the optimized procedures.