Biobanking Science: Biobank Sperm Without Liquid Nitrogen

New research suggests that biobanks can freeze, transport and store sperm for up to two years at -80°C without compromising quality.

Liquid nitrogen free sperm biobanking
It may be possible to biobank mouse sperm without using liquid nitrogen.
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With gene editing technology such as CRISPR, it is becoming quicker and easier to generate disease models for medical research. Researchers around the world use tens of thousands of different mouse lines. Research institutions can save money and reduce unnecessary use of animals if they biobank any mouse lines that are not actively used. It is also more humane to transport frozen embryos or sperm rather than ship live animals.

At the moment, it is more common to biobank embryos, although this may change as sperm is more resistant to freeze-thaw damage than embryos. Sperm is also technically easier to freeze and biobank than embryos. One caveat with biobanking sperm is that it requires in vitro fertilization to regenerate the biobanked mouse line.

The gold standard biobank method for both embryos and sperm is cryopreservation in liquid nitrogen. However, liquid nitrogen systems can be expensive to set up and maintain. Therefore, many biobanks, laboratories and clinics use a combination of liquid nitrogen freezers/dewars and -80°C freezers to store samples. A biobank that stores mouse lines as sperm may be able to switch to using only -80°C according to new research from the European Mouse Mutant Archive (EMMA). EMMA is a central biorepository and network of biobanks across Europe that preserves and distributes mouse lines for research.

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A 2017 study by EMMA researchers found that mouse sperm maintains quality and function when transported on dry ice for up to seven days (1). This means that biobanks and laboratories could share mouse lines by shipping sperm instead of sending live animals. It is cheaper and easier to ship biological materials on dry ice than in liquid nitrogen. The same research group also found that mouse sperm can be stored in -80°C freezers for up to two years without affecting viability (2).

In a third study published in 2018, EMMA researchers directly compared sperm quality after cryopreservation via liquid nitrogen freezing or slower freezing using a -80°C freezer. They used common cryoprotectant CPA with monothioglycerol or L-Glutamine, and stored samples for up to one year before assessing sperm quality. The researchers found that sperm had the same viability and fertility with both freezing methods. They assessed viability by thawing the sperm and using it for in vitro fertilization. The same number of live mouse pups were born from sperm frozen by liquid nitrogen as from sperm frozen in a -80°C freezer. The researchers also found that mouse sperm could be transferred from a -80°C freezer to liquid nitrogen without adversely affecting quality and function.

The results of these three studies imply that biobanks may not need to use liquid nitrogen to freeze and store mouse sperm. However, this research only looked at the effect of -80°C storage for up to two years. Also they only used sperm from a small number of mouse lines. Since biobanks will likely need to store many mouse lines for much longer than two years, more research should be done looking at the effect of -80°C storage over longer time periods and should include sperm from a wider selection of mouse lines.

Also this finding does not apply to embryos or other cell lines that have been shown to deteriorate if stored at -80°C rather than cryopreserved in liquid nitrogen.



1. Raspa et al. Dry ice is a reliable substrate for the distribution of frozen mouse spermatozoa: A multi-centric study. Theriogenology. 2017
2. Raspa et al. Long term maintenance of frozen mouse spermatozoa at -80°C. Theriogenology. 2018
3. Raspa et al. A new, simple and efficient liquid nitrogen free method to cryopreserve mouse spermatozoa at -80°C. Theriogenology. 2018