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Why Is Liquid Nitrogen Used for Cryopreservation?

Liquid nitrogen is a coolant used in cryopreservation. What makes it good for long-term storage?

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Nitrogen is the most abundant element in earth's atmosphere. That alone could act as a sufficient answer for some as to why we use liquid nitrogen for cryopreservation. But of course it isn’t quite as easy as “grabbing nitrogen” out of thin air. Abundance alone doesn’t make a coolant fit for storing Tomorrow patients for an indefinite time frame. In this article we will highlight what really makes nitrogen, in its liquid form, the best option for biostasis today.

How does cryopreservation work?

The cryoprotection process start by slowly lowering the core temperature of the body to sub-freezing levels. This puts it below the so-called “glass transition temperature” at around -130°C achieving vitrification in the process. Cryoprotective agents are used to protect the cell structure from hazards like ice-crystal formation. Through cooling and the cryopreservation process, metabolic rates are reduced significantly, stopping biological activity completely. Once vitrified, the body is in complete “biological pause”, meaning that patients can be stored in this state indefinitely without degradation.

At Tomorrow Bio, we use temperatures of -196°C for long-term storage of cryopreserved patients (with Intermediate Temperature Storage aka ITS in development), which is achieved by utilising liquid nitrogen.

What is Liquid Nitrogen?

Liquid nitrogen is a colorless, odorless, and tasteless gas with a boiling point of -196°C and a freezing point of -210°C. It is generated on a large scale in air separation plants. 

Here, air is split into its primary components (mostly nitrogen and oxygen) by compressing, purifying and ultimately cooling it close to its liquefaction temperature.

Through pressurized super-insulated vacuum vessels, these split components can be stored (for many weeks) and transported to wherever they might find good usage. Amongst other things, liquid nitrogen is being put to use at cryopreservation storages inside of cryogenic storage dewars.

A donut chart of composition of dry atmospheric air
One thing is for certain: We will not run out of liquid nitrogen in the future.

Why Is Liquid Nitrogen Used for Cryopreservation?

Liquid nitrogen is commonly used to cool substances to extremely low temperatures. It’s fortunate that it also possesses a natural temperature below the glass-transition point, which cryopreservation procedures can work with. 

Moreover, it is relatively cheap, especially compared to similar gasses (e.g. liquid neon). Because liquid nitrogen can be extracted from ambient air, it is an environmentally friendly gas as well - an important factor when regarding the long storage time it is supposed to enable. The substance is also exceptionally easy to sustain. A single batch of it is able to keep cryogenically stored patients in optimal conditions for more than a week. At the top of the dewars, liquid nitrogen constantly boils off as it evaporates into gas. To counteract this, the storage facility gets supplied with liquid nitrogen on a weekly basis, so that after a week new liquid nitrogen is simply added to keep the temperature stable. 

In worst-case scenarios, where refilling is impossible for some reason, it is able to protect the brain of a patient for up to a month without issue. This is due to our patients being stored upside down within the dewars, allowing for the brain to benefit from the best protection. In the very unlikely case of accidental thawing, the brain would be the very last thing to be exposed.

Thanks to advanced storage technology, we’re able to keep additional stock as a backup, so even if the supply chain runs into unforeseen trouble, we are prepared.

How Is Liquid Nitrogen Used?

Whole-body cryogenic storage dewars are filled up to the top with liquid nitrogen, until it submerges the patient's body entirely. Refilling occurs as we mentioned before.

In less common ITS dewars, only the bottom gets filled with the substance. This approach allows for the body to be stored at a higher temperature, reducing the chance of thermal damages. One drawback of ITS, however, is that it has to be refilled more frequently, about once every few days. 

Replenishing can be conducted automatically, but is usually done by hand so as to not be dependent on electricity. This lowers the overall cost of storage drastically over time and makes the upkeep more sustainable to boot.

 

Alternative Options (and why they aren’t used)

Despite all the praise we’ve given liquid nitrogen so far, it is not perfect. For one, its temperature and the glass transition temperature are more than 60°C apart, around -196°C and -130°C respectively. Liquid nitrogen’s natural temperature works with cryonic procedures and is the easiest to uphold, which is a key aspect of why we are using it. However, ideally we would want to stay as close to the glass transition point as possible, to prevent thermal stress due to cooling further. ITS Storage systems are trying to solve this issue, but come with their own added challenges and extra cost.

There are additional substances that could be used instead of liquid nitrogen but, for one reason or another, they aren’t. Let’s take a look at some of them:

Liquid Hydrogen (LH2)

  • Decent price 
  • Expensive equipment needed for handling
  • Very low boiling point of -252°C (remember, we don’t want to go too far below 130°C)
  • Evaporates too fast
  • Highly flammable

Liquid Neon(LNe)

  • Completely inert (unaffected by other chemicals)
  • Too cold (-246°C)
  • Extremely expensive! (€120+ per liter if not more)

Liquid Oxygen(LO2,LOX)

  • Cheap
  • Good oxidizer (can burn most organic matter and also metals, you and the entire facility)

Liquid Helium(LHe)

  • Can freeze at very low pressures
  • The most inert substance known
  • Even lower temperature (-269°C)
  • Special dewars needed
  • Evaporates extremely easily
  • Predicted to run out in a few decades

Below the line, all of these options are either too expensive, too cold or too unsafe to make sense. In comparison, liquid nitrogen easily beats them all.

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Conclusion

Liquid nitrogen remains the best option for now. As long as no new substance is discovered that works exceptionally well for cryopreservation procedures, it is more likely that new techniques and technologies using liquid nitrogen will improve storage quality instead (like ITS).

It’s the cheapest, and easily sustainable way to uphold Biostasis for humans we know. Other options exist, but don’t make much sense in comparison to liquid nitrogen. Looking towards the future, we will most likely see this natural coolant used in cryopreservation for a long time to come.

Interested in finding out more about biostasis aka. cryonics? Feel free to schedule a call with us any time! We are happy to talk to you and answer all your questions!

Tomorrow Bio is the worlds fastest growing human cryopreservation provider. Our all inclusive cryopreservation plans start at just 45€ per month. Learn more here.
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