There exists a theoretical possibility of contamination of samples and embryos during liquid nitrogen storage. This risk is greatest for tubes that are submerged in liquid nitrogen rather than in the vapour phase. If viable microorganisms were present in the liquid nitrogen then improperly sealed tubes would be at risk. Contamination could also occur post retrieval if the vials were not properly disinfected before placing in the tissue culture hood. Under proper operating conditions it is usually considered that such risks are negligible.
A new study led by Francisco Marco-Jimenez of Universitat Politècnica de València, Spain, aimed to determine if such assumptions of negligible risk were valid during storage of rabbit embryos in a liquid nitrogen biobank. The research team evaluated the risk of cross-infection of sterile embryos from contaminated liquid nitrogen, and cross-contamination of sterile liquid nitrogen from infected embryos, using naked (open topped) and closed devices stored for one year. The results were published in the journal Animals.
When purposefully bacterially contaminated liquid nitrogen was used in tanks to store rabbit embryos it was found that open storage tubes that were not previously infected became contaminated with viable microorganisms. Tubes that were closed and sealed did not become infected.
When purposefully infected embryos in open-topped tubes were placed in sterile liquid nitrogen tanks it was found that the liquid nitrogen became contaminated with viable organisms, however the concentration was apparently not sufficient to infect any other non-infected vials; open or closed.
The study therefore suggests that under proper storage conditions the risk of embryo cross-contamination is indeed negligible, unless the liquid nitrogen itself is highly contaminated. Testing the sterility of liquid nitrogen in storage tanks may have some merit under certain circumstances and depending on the tubes used for storage. That said, using sealed tubes and storing in the vapour phase could negate the risk.
“Our findings clearly support that both cross-contamination and cross-infection during embryo storage in liquid nitrogen biobanks are plausible. So, to ensure biosafety for cryogenic storage, the use of closed systems that avoid direct contact with liquid nitrogen must be considered. Moreover, it seems essential to provide best practice guidelines for the cryogenic preservation and storage of gametes and embryos, in order to define appropriate quality and risk management procedures,” concluded the authors.
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