CryoClean technology: A novel closed vitrification system combining high efficiency and sterility for embryo...

Area of Science:

• Assisted Reproductive Technology (ART)
• Cryobiology
• Embryology

Background:

• Traditional open embryo vitrification systems risk contamination, while closed systems compromise cooling efficiency.
• Bridging the safety-efficiency gap in embryo cryopreservation is crucial for clinical assisted reproductive technology.

Purpose of the Study:

• To introduce and evaluate CryoClean, an innovative closed vitrification technology using sterile liquid air.
• To assess the cooling efficiency, biosafety, and developmental outcomes of CryoClean compared to existing methods.

Main Methods:

• CryoClean utilizes a "freeze-first, seal-later" principle with sterile liquid air (≈14.5% O2) as a refrigerant.
• Cooling rates were compared to direct liquid nitrogen immersion; microbial cultures assessed refrigerant sterility.
• Mouse 8-cell embryos were vitrified using CryoClean and Cryotop (open system) for comparative analysis of survival, blastocyst formation, and hatching rates.

Main Results:

• CryoClean achieved cooling rates equivalent to direct liquid nitrogen immersion.
• Sterile liquid air confirmed absence of microbial contamination, unlike stored liquid nitrogen.
• CryoClean demonstrated comparable post-thaw survival (100%), blastocyst formation (84.8%), and hatching rates (64.3%) to the open Cryotop method.
• Whole-genome bisulfite sequencing showed similar hypomethylation patterns between CryoClean and Cryotop groups, with no significant epigenetic differences from fresh embryos.
• Offspring from CryoClean-vitrified embryos exhibited normal postnatal development, organ weights, serum biochemistry, cardiac function, and reproductive capacity.

Conclusions:

• CryoClean effectively combines the high efficiency of open vitrification systems with the biosafety of closed systems.
• This technology presents a promising advancement for sterile embryo vitrification in clinical assisted reproductive technology.
• CryoClean overcomes the efficiency bottleneck in sterile embryo cryopreservation, ensuring safety and developmental competence.