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When biological samples are stored at cryogenic temperatures using liquid nitrogen there are two primary options to consider. The first is vapor phase storage and the second is storing samples directly in liquid phase nitrogen.
First used in the 1950s for cryogenic storage, liquid nitrogen has made it possible to store samples at temperatures down to -196 °C. Since then, LN2 has been deemed to be the benchmark where cryogenic storage is concerned. Therefore, a cryogen like liquid nitrogen is the only solution for the long-term storage of samples below the glass transition point of water (-135 °C), the point at which virtually all biological functions stop.
Initially, the only option available was to submerge the sample containers in liquid phase. However, progress in the technology led to the emergence of dry or vapor phase storage - where samples are stored above the LN2 rather than in it.
Both kinds of sample storage freezer design are still in use today, each offering a unique set of pros and cons. Generally, the most optimal choice will rely on individual needs and requirements.
Storage is where the advantages and disadvantages of each solution are most clear. Due to the differing designs of freezer interior, both come with their own benefits and drawbacks.
Liquid phase nitrogen is present in both types of freezer. Rather than being stored directly in the liquid nitrogen, however, vapor phase freezers store samples in the vapor created above. There are those that criticise vapor phase storage freezers due to the lack of space: the lower third of the freezer is left unusable due to the requirements for the liquid nitrogen base.
However, this is now an outdated criticism as it largely refers to liquid phase freezers which were simply modified for nitrogen vapor storage. Newer units designed specifically for the purpose don’t tend to experience such storage limitations.
One of the principal features in the design of vapor phase cryogenic freezers that most users can agree on is that the chances of contamination are minimised, a significant issue concerning liquid phase LN2 freezers. Moreover, vial explosion is another potential issue that is avoided in vapor phase storage but is a phenomenon that can take place when just a single drop of LN2 penetrates a sample vial. Subsequently, when heated, vials can wildly explode due to the prompt expansion of the liquid nitrogen to 690 times its volume. This phenomenon is not only potentially dangerous, but would also result in sample loss.
If stored improperly in liquid nitrogen there is a chance contamination can occur in samples. Thus, precious samples should not be stored with potentially infectious samples in liquid phase LN2 for this very reason.
Even when suspended in liquid nitrogen, some contaminants, including Hepatitis B and Vesicular Stomatitis Virus, continue to be extremely infectious. One report claimed that Hepatitis B retained its infectivity after two years in liquid nitrogen.
Vapor phase LN2 freezers bypass contamination issues as samples are stored above the LN2 rather than in it, reducing the potential of transmission via the liquid itself.
Additionally, researchers can be at risk of contagion due to the contamination of the outer layer of the containers during retrieval. Unlike liquid phase freezers, there is a much lower-risk of user contamination or injury from splash-back with vapor phase freezers as there is no contact required with the liquid nitrogen itself. Disregarding the contamination possibilities, LN2 drag out on its own can raise enough of a threat, not only to operators but also the surrounding environment.
For longer term sample storage, liquid phase LN2 freezers are typically preferred as submersion directly in the liquid is deemed to be more reliable and efficient over an extended period of time.
In contrast, freezers developed specifically for vapor storage have an advantage when it comes to accessing samples. This is due to the fact that it is much easier to retrieve samples from vapor suspension. For example, the HEco vapor phase freezer enables easy access to the samples via the lid and great storage density without compromising hold time or sample security.
The HEco also includes an internal carousel that transports the sample to the operator, eradicating the struggle of reaching samples at the back of the freezer - a real hazard when using liquid phase freezers.
Some believe that liquid storage consumes less LN2. However, high performance, efficient purpose-built vapor storage freezers, such as the HEco, consume nearly 50 percent less liquid nitrogen than the equivalent capacity liquid storage freezer.
Whilst for several years researchers were usually restricted when it came to temperature control, the introduction of vapor phase freezers initiated a more effective temperature adjustment.
Historically, operators have been skeptical when it comes to temperature uniformity in vapor phase freezers as early versions of the technology presented significant differences between the temperatures of the samples at the top and bottom of the freezer. However, these initial shortcomings have been overcome in contemporary vapor phase freezers: today’s versions are able to comfortably hold a temperature of -190 °C in the top box. This is no mean feat, especially when the consideration is made that a modern vapor phase freezer such as the HEco can sustain cryogenic temperatures for several weeks even after removal of the LN2 source.
Liquid phase freezers are limited to operation at a predetermined temperature of around -196°C. Whereas innovations and advancements in vapor storage, such as the Variō freezer from MVE Biological Solutions, now provide the opportunity for temperature control. The Variō stores liquid nitrogen in a separate chamber which is then driven through a heat exchanger to create vapor. Consequently, this novel freezer has the capacity to effectively operate across a broad temperature range; from -20 °C to -150 °C.
Dependent on the conditions and needs of a cryo-room or facility, the decision between liquid and vapor phase freezers is not always an obvious one as any judgment will need to consider an assortment of factors including temperature, accessibility, and storage needs.
About Air Products PLC
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