How two simple tips can prevent your cryogenic vials from leaking (or exploding)

Two simple rules to prevent leakage of your vital samples: Secure the cap seal and do not overfill the vial.

With ultra-low temperature (ULT) sample storage, one of the most preventable ways that precious samples get damaged or lost occurs when excess pressure builds up inside of a cryogenic vial or glass ampule during freeze/thaw cycles.

Although the cap seal on some vials surpass the IATA minimum standard (15 PSI) for internal pressure, the initial and most easily prevented cause is always the same: exceeding a cryogenic vial’s critical fill volume.

 

Take a 2 mL cryogenic vial containing tissue culture media and DMSO (10% DMSO, a common cryoprotectant for animal cells) as an example. What goes on inside the vial at extremely low temperatures?

The media and DMSO expand when frozen to -80 or -196C, compressing the gas contained in the area above the liquid level. If the 2 mL vial is properly filled to the recommended critical fill volume of 1.8 mL, the resultant change in volume and pressure shouldn’t increase above tested levels and the cap seal is not compromised.

However, if a 2 mL vial is filled with 2 mL of sample, the change in volume and pressure (from 2 ml to 2.12 ml, with a 2 fold pressure increase or to levels greater than 36 PSI) may be enough to compromise the cap seal for some cryovials, inviting contaminants, or worse, liquid nitrogen (LN2) into the vial. If other reagents, such as glycerol, ethanol or methanol are used in the freezing media, the pressure changes can be even greater.

When even a small amount of LN2 enters a cryovial, a small explosion is waiting to happen when it is taken out of a freezer. LN2 has a liquid to gas expansion ratio of 1:694, which generates a tremendous amount of force on a cryogenic vial cap seal.

In other words… POP!

So, the take home message here is that you can safely store your vital samples every time, with great consistency, as long as you secure that cap seal. Tighten the cap, and remember the simple rule of not exceeding a cryogenic vial’s critical fill volume.

Key Points:
1. Tighten cap prior to freezing.
2. Do not over fill containers.
3. Before thawing, loosen cap under laminar flow hood to allow pressure to release. Retighten prior to placing in water bath!
4. LN2 is EXTREMELY COLD and can cause SEVERE burns or damage to skin and eyes. USE CRYOPROTECTIVE GLOVES AND PROTECTIVE EYE/FACESHIELD.

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