Achieving Container Closure Integrity at Cryogenic Storage Temperatures

Gene and cell therapies stored at cryogenic temperatures, from -150°C to -180°C, place demands on your vial stopper combination that traditional cold-chain packaging was never designed for. Container closure integrity (CCI) can be lost at these extremely low temperatures and regained once the vial returns to room temperature — a transient failure that only laser-based headspace analysis is able to detect for large sample sets.

A recent LIGHTHOUSE case study investigated primary packaging for cryogenic storage using a 2 mL hybrid vial from SiO2 Materials Science, a polymer container with a microscopic, glass-like barrier coating on the inside surface. This was tested against a standard 2R borosilicate glass vial. Both vial types were sealed with the same stopper, a readily available West 4432 chlorobutyl elastomer.

Because the quality of the capping process affects CCI outcomes, stoppered samples were prepared across four capping pressure groups, from loosely to tightly capped, based on residual seal force (RSF) measurements. Matched sample sets of both vial types were stored in a liquid-nitrogen-cooled cryogenic freezer for one week. After storage, all samples were measured CCI using non-destructive laser-based headspace analysis.

The results were consistent across every capping pressure group. Every 2R borosilicate glass vial, regardless of how tightly it was capped, lost container closure integrity during cryogenic storage. Every SiO2 hybrid vial, across the same four capping groups, maintained it.

This builds on earlier published work showing that particular vial-stopper combinations can deliver good CCI performance at -80°C when the vial sealing process is well controlled. Very few packaging systems, however, have demonstrated acceptable CCI performance at true cryogenic temperatures. The SiO2 hybrid vial, sealed with a standard chlorobutyl stopper, is a promising candidate for those storage conditions.

Testing packaging at -150°C to -180°C requires a method that can catch a failure mode that disappears once the vial warms back up. The case study used a laser-based headspace method to measure oxygen depletion. The cryogenic freezer used the gaseous phase of liquid nitrogen to reach temperature, creating a nitrogen-rich storage environment. A vial that loses CCI during storage ingresses nitrogen through the leak path, displacing the oxygen in its original air headspace. Measuring that oxygen depletion after storage identifies which vials failed, even though the leak path itself has closed again by the time the vial is tested.

The method is deterministic, non-destructive, and analytical. In this study it detected everything from gross leaks down to micron-sized defects, giving confidence in its sensitivity across the range that matters for cryogenic packaging qualification.

Bringing a product to market that requires cryogenic storage means generating analytical data that your primary package can hold CCI at storage temperature, and validating that it holds through the transport chain as well. That starts with a CCI test method capable of catching transient failures, a method such as laser-based headspace analysis.

If your product requires cryogenic storage, the underlying question is which container closure system and capping process can retain CCI at your target temperature, and more importantly, how you generate robust data to prove this to the regulator.

Talk to a LIGHTHOUSE packaging expert to discuss CCI testing for your cryogenic storage program, or get the full case study to see how we performed this particular study.