An auto-nucleating device includes a tube containing a crystalline cholesterol matrix. The ends of the tube are closed by a membrane impermeable to the cholesterol but permeable to liquids contained in a cryopreservation vessel. The auto-nucleating device provides a site for ice nucleation during freezing of the liquid within the vessel. The cryopreservation vessel can be a flexible vial having a closed port adapted to be pierced by a needle, and an opposite end that is initially open to receive the liquid. Another vessel includes an adaptor mounted to liquid container with a tubular branch closed by a needle septum and another tubular branch provided with a barbed fitting for engaging a flexible tube that terminates in a needle septum. Alternatively, the vessel includes an inlet and vent branch at the top of the container and an outlet septum at a bottom opening.
